第十三届全国中药与天然药物药理学术会议论文摘要

2021年10月29-31日，陕西 咸阳

为促进中药药理学科的学术交流与发展，提高中药药理学及中药新药研发水平，由中国药理学会中药与天然药物药理专业委员会与陕西中医药大学联合举办的“第十三届全国中药与天然药物药理学术交流会”于2021年10月29-31日在陕西省咸阳市举行。本次会议展示我国中药、天然药物和传统药物的基础药理学和临床药理学的最新研究进展，研讨热点问题，旨在促进交流与合作，提高研究水平。

会议议题为中药和天然药物的基础药理学、临床药理学和毒理学，基于现代生物医药科学的新理念、新策略和新技术的中药和天然药物药理学研究，中药和天然药物的新药研究与开发和中药和天然药物抗重大传染病研究。会议主要包括三方面内容：一是学术交流，形式为大会报告、专题报告、青年学术论坛及壁报展讲；二是举办“中医药治未病相关药理学研究及新药研发”专题研讨会，针对《中医药发展战略规划纲要（2016-2030年）》提出的中医药在治未病中的主导作用是未来中医药发展的战略定位之一，对相关重大科学和技术问题进行研讨，为提高当代健康理念及重大疾病防控水平提供参考（全体与会代表均可参加）；三是召开中国药理学会中药与天然药物药理专业委员会会议。学术交流形式为大会报告、专题报告、青年学术论坛和壁报展讲。

会议报告

Progress in anti-inflammatory effects of escin

FU Feng-hua1,2,WANG Tian1,2

（1.School of Pharmacy,2.Key Laboratory of Molecular Pharmacology and Drug Evaluation of Ministry of Education,Yantai University,Yantai 264005,China）

Abstract：The fraction of horse chestnut seeds was named escins,which mainly consists of A,B,C,and D escin.Accumulating evidence suggests that escin exerts potent anti-inflammatory and anti-edematous effects.The effects of escin on inflammation and edema have been confirmed in various models.In a study in 1961,intravenous administration of escin was found to reduce acute edema in a rat paw.In the same study,escin was found to inhibit the increase in vascular permeability induced by egg white injection.Escin dose-dependently reduced the capillary permeability in chlo⁃roform-induced local inflammation in the abdominal skin surface of rabbits.The anti-inflammatory and anti-edematous effects of external use of escin were studied in carrageenan-induced paw edema and histamine-induced capillary perme⁃ability in rats.Escin gel decreased the contents of PGE2,TNF-α,and IL-1β,and reduced the raw edema and capillary permeability.The carrageenan-induced paw edema and pleuritis in bilaterally adrenalectomized rats were used to inves⁃tigate the anti-inflammatory effects of escin and glucocorticoid alone or combined.Co-administration of escin with corti⁃sone significantly reduced the volume of exudates and the number of white blood cells of exudates.The findings sug⁃gest escin can synergize with glucocorticoids to enhance their anti-inflammatory effect.The anti-inflammatory effect of escin was investigated in carrageenan-induced paw edema and acetic acid-induced capillary permeability in mice.Escin showed an anti-inflammatory effect,which is similar to that seen with dexamethasone treatment.However,escin showed a longer duration of the anti-inflammatory response than that of dexamethasone.Furthermore,escin had no signif⁃icant effects on spleen index,thymus index,proliferative capacity of splenocytes,lymphocyte count,and phagocytic rate.The findings suggest that escin is a potent anti-inflammatory drug with long-lasting anti-inflammatory effects without any immunosuppressive effects.Traditionally the mechanism of anti-inflammatory effect of escin is supposed to be rela⁃tive to release of PGF2α and corticosterone.The early studies showed that escin might promote the release of PGF2α and affect the pituitary adrenal system,stimulate the release of adrenocorticotropic hormone（ACTH）and glucocorticoid,which may explain its anti-inflammatory and anti-edema effects.Escin has glucocorticoid-like anti-inflammatory effect.However,escin did not exhibit an anti-inflammatory effect in low dose.Combination of suboptimal concentrations of escin with corticosterone inhibited the release of inflammatory factors including NO,TNF-α and IL-1β in the LPS-stimulated macrophage cells.Previous studies demonstrate that escin combined with glucocorticoid produced synergistic antiinflammatory effects.The potential synergistic mechanisms may be associated with the property which escin regulates the glucocorticoid receptor（GR）signaling pathway.Escin can upregulate the expression of GR,promote the combina⁃tion of glucocorticoid and GR,then promote the activated GR transfer into the nucleus.Activated GR will inhibit the acti⁃vation of NF-κB directly,thus further inhibiting the expression of TNF-α and IL-1β and other inflammatory factors.Escin could inhibit 11β-HSD2 but not 11β-HSD1,thus decrease the metabolism of glucocorticoid.Escin and glucocorticoids have similar chemical structures.This indicate that one of the anti-inflammatory mechanisms of escin may be due to its stimulating GR by binding to it.Eacin might be a partial agonist of GR.A good many of researches have demonstrated the anti-inflammatory properties of escin,and shed light on the underlying mechanisms by which escin exerts these effects.Escin,as an oral or intravenous formulation,or a topical gel,inhibits inflammation,producing measurable improve⁃ments in edema and acute lung injury.Further clinical studies of escin are needed to demonstrate these properties in larger patient populations.

Key words：escin;inflammation;edema;capillary permeability;acute lung injury

Corresponding author：FU Feng-hua,E-mail：fufh@ytu.edu.cn

Naringenin prevented nonalcoholic steatohepatitis fibrosis via regulating MAPK/FoxO3a pathway and promoting apoptosis of activated hepatic stellate cells

YUE Shan-shan,QI Rong

（Department of Pharmacology,School of Basic Medical Sciences,Peking University Health Science Center,Beijing 100191,China）

Abstract：OBJECTIVEThe pathological characteristics of nonalcoholic steatohepatitis（NASH）include liver steato⁃sis,inflammation,and fibrosis.Fibrosis is the most severe and significant pathological feature in NASH.Effective drug treatment could reverse early liver fibrosis and is of significance to prevent NASH from progressing into cirrhosis and liver cancer.Identification of drug targets for NASH treatment has been an active research area and is essential for the development of anti-NASH medications.Naringenin（NGN）is a flavonoid compound rich in citrus fruits.Our preliminary data demonstrated that NGN reduced diet-induced lipid accumulation and inflammation in the mouse liver,but whether NGN can attenuate liver fibrosis of NASH is not known.METHODSTo study the effect of NGN on NASH fibrosis.WT mice were fed with high fat diet（HFD）and injected intraperitoneally 20%carbon tetrachloride at the same time for 8 weeks to induce NASH,and NGN was administrated by gavage in the meantime.In vitro,LO2 cells and LX2 cells were stimulated by oleic acid（OA）combined with lipopolysaccharide（LPS）,respectively.RESULTSTreating the WT mice with NGN 100 mg·kg-1·d-1significantly attenuated hepatic lipid accumulation,hepatic fibrosis,plasma ALT and AST levels,inhibited protein expression of p-ERK,p-FoxO3a in the mouse livers.In vitro,on OA and LPS stimulated LO2 or LX2 cells,NGN significantly promoted apoptosis of activated hepatic stellate cells while inhibited apoptosis of hepatocytes.Mechanism study indicated that NGN inhibited MAPK pathway and promoted activation of FoxO3a,conse⁃quently promoted apoptosis of the activated LX2 cells and inhibited liver fibrosis.CONCLUSIONNGN preventes NASH fibrosis via regulating MAPK/FoxO3a pathway,thus promoting apoptosis of the activated hepatic stellate cells.

Key words：nonalcoholic steatohepatitis;liver fibrosis;hepatic stellate cells;apoptosis;MAPK;FoxO3a

Corresponding author：QI Rong,E-mail：qirong0311@163.com

近十年中国国家自然科学基金会中药药理学与药物药理学申请和资助情况比较分析

毕明刚

（中国国家自然科学基金委员会，北京 100085）

摘要：从中国国家自然科学基金委数据库采集2012-2021年中药药理学与药物药理学面上项目、青年科学基金项目和地区科学基金项目的申请和资助项目相关信息进行回顾性分析，比较两者在相关领域的差异。结果显示，在申请量上药物药理学与中药药理学基本持平，在资助数量和资助率上药物药理学高于中药药理学；在新技术、新方法、研究热点等关键词方面，药物药理学早于中药药理学数年出现；在研究对象上，药物药理学多集中于单一化合物，中药药理学尚有一定比例的复方或活性部位；在研究思路上，药物药理学更集中于某一作用途径的深入研究，中药药理学多倾向于多途径、多靶点的研究。中国国家自然科学基金的资助促进了中药药理学的发展、学科的建设和人才的培养，同时也存在对中医药领域的支持力度相对不足、学科发展不平衡、研究成果缺乏连贯性和延续性、优势病种局限、科研思路过于仿效国际热点、高精尖人才相对缺少等不足。建议应针对存在的问题，扬长避短，以临床科学问题为导向有针对性的研究。

关键词：中国国家自然科学基金；中药药理学；药物药理学；比较分析

通讯作者：毕明刚，E-mail：bimg@nsfc.gov.cn

经典方剂二至丸补肝益肾作用研究

毛浩萍1，2，戴永娜1，2，陶 蕊1，2，韩立峰1，2，杨文志1，2，高秀梅1，2

（天津中医药大学1.中医药研究院，2.教育部方剂学重点实验室，天津 301617）

摘要：目的二至丸由墨旱莲、女贞子2味药配伍而成，具有补腰膝、壮筋骨、强肝肾、乌髭发的功效，是补益肝肾的经典代表方。本研究以二至丸为研究对象，以雌雄激素样作用为切入点，阐明二至丸体内药效物质基础，揭示二至丸治疗肝肾阴虚证的化学活性物质和生物学基础，为临床精确使用二至丸提供科学依据，诠释中医“异病同治”科学内涵。方法采用LC-MS、LC-NMR-MS等技术对分离得到的二至丸中化合物进行鉴定，明确二至丸口服吸收入血的化学物质及药动学特征；基于高内涵筛选及报告基因技术确认二至丸及入血成分雌雄激素样活性；利用与肝肾阴虚症状表现类似的动物模型，围绕骨、脂肪及前列腺等组织研究二至丸中雌雄激素样活性成分的作用特点及靶点。结果①基于UHPLC/Q-Orbitrap-MS建立一种同时靶向与非靶向扫描技术，结合离线全二维色谱，从二至丸中在线分离监测7大类共270个化学成分。研究了墨旱莲中7种入血成分的体内代谢过程，结果发现其均符合一室开放药动学模型，大部分在1 h左右血药浓度达到峰值。同时其入血成分最终鉴定4大类共85个代谢产物。②对272个来源于包含二至丸的化合物进行筛选，发现33个化合物在10 μmol·L-1浓度下明显促进雌雄激素受体核转位，其中山奈酚、齐墩果酸具有选择性雄激素受体调节剂样作用。③二至丸具有抗骨质疏松作用，其作用的分子机制与降低破骨细胞及脂肪细胞分化；升高成骨细胞分化以及调控肠道菌群有关。④二至丸抑制雌激素不足导致的肥胖。通过AMPK信号通路调节细胞能量稳态平衡，并且作用于雌激素受体以间接调节过氧化物酶体增殖物激活受体γ辅激活因子1α（PGC-1α），从而干预线粒体结构与功能，调节适应性产热，参与脂代谢。⑤二至丸抑制良性前列腺增生（BPH）。二至丸显著减少BPH平滑肌细胞层厚度，下调PCNA表达，显著增加ERβ和MMP-1的表达，有效降低睾酮、E2、DHT、AR、前列腺特异抗原（PSA）、ERα、SRD5A1/2、芳香化酶、p-ERK、α-SMA、collagen Ⅰ、TGF-β1和p-Smad3的表达，缓解BP的生物学进程。结论补益肝肾经典方剂二至丸及其成分具有植物雌雄激素样作用；二至丸的植物雌雄激素样作用是其抗绝经期骨质疏松症、绝经期激素不足导致的肥胖症及抑制良性前列腺增生等作用的生物学基础。

关键词：二至丸；补肝益肾；植物雌雄激素

通讯作者：高秀梅，E-mail：gaoxiumei@tjutcm.edu.cn

基于中医“治未病”理论的芪珀生脉颗粒治疗房颤的应用研究

胡元会，王 欢

（中国中医科学院广安门医院，北京 100053）

摘要：“治未病”思想最早记载于《内经》，是中医学理论体系的重要内容之一。在临床实践中，我们长期致力于心血管疾的研究，其中研究显示芪珀生脉颗粒以益气养阴、祛风安神为治法，通过控制危险因素、控制房颤发展，从而达到未病先防、已病防变、瘥后防复的作用。

我国古代哲学思想对“治未病”理念形成有深远影响，“治未病”肇始于《素问·四气调神大论篇》，后世医家经过不断创新和发展，提出了以“未病先防，已病防变，瘥后防复”为核心的中医“治未病”理论体系。体现中医治病“防重于治”，强调人不仅自身是一个有机整体，还应遵循人与自然、社会环境的和谐统一，即“天人合一”整体观。

中药芪珀生脉颗粒治疗房颤的核心在于中医“治未病”理念，重点在于“未病先防，既病防变，瘥后防复”3个环节。房颤的病机实质为本虚标实，尤以本虚为关键。其中气阴两虚证是阵发性房颤的主要证型之一，芪珀生脉颗粒治疗气阴两虚型房颤疗效显著。①未病先防：在房颤未发生前，机体往往产生相关的病理改变，如合并房性心律失常房早、房速、左房结构改变等人群发生房颤的风险较高，中医药及早适时干预，是预防房颤发病的重要措施。2010年欧洲心脏病学会心房颤动管理指南提出“上游治疗”的新理念，目的是在房颤发生前从致病因素层面防止疾病发生及进展。研究显示，芪珀生脉颗粒可以通过减轻炎症、氧化应激和纤维化等机制改善心房基质重构，改善房颤发生的“上游机制”。②既病防变：对已经发生房颤的病人，强调重在减少或终止房颤发作、预防合并症的发生。研究提示芪珀生脉颗粒可抑制血小板的活化，降低血栓风险，同时可有效缩短房颤的持续时间，延长心房有效不应期，防止向持续性房颤发展。③瘥后防复：对于房颤患者，延长窦性心律的时间是防止房颤复发的关键因素。房颤发生的直接因素是电传导的异常，而结构基础是心房结构的异常。芪珀生脉颗粒可以改善房颤患者心房电重构和结构重构，还可以通过优化心房脂肪酸和葡萄糖的利用，促进线粒体生物合成，拮抗线粒体功能障碍引起的房颤。中医“治未病”可谓是中医的预防医学，芪珀生脉颗粒即是中西医结合治疗房颤在“治未病”理念的生动体现，“治未病”观在现代医学的防治过程中有重要指导作用和应用价值。

关键词：中医；治未病；芪珀生脉颗粒；房颤

通讯作者：胡元会,E-mail：huiyuhui55@sohu.com

基于天然产物的药理活性成分发现与作用机制研究

李静雅

（中国科学院上海药物研究所，上海 201203）

摘要：代谢综合征是以机体的蛋白质、脂肪和碳水化合物等代谢紊乱为特征的病理状态，是导致糖尿病、代谢性脂肪肝病和急慢性肾病等的危险因素。线粒体是细胞内提供ATP的能量工厂，也是三大类物质代谢的中心枢纽。大量临床与基础医学研究均表明，线粒体氧化呼吸活力下降、线粒体定位蛋白遗传突变或转录下调等，与肝实质细胞凋亡、脂肪组织产热、胰腺β细胞分泌胰岛素以及骨骼肌胰岛素抵抗等息息相关。基于外周组织（包括肝、肾、胰和脂肪组织）功能性实质细胞的线粒体功能紊乱，研究团队建立相关的功能表型模型，展开药食同源以及传统中药民族药来源的天然产物活性成分筛选，确认药理活性并探索其作用机制。近年来发现，线粒体自噬促进剂硫黄菊素（sulfuretin）保护肝细胞凋亡（Acta Pharmacological Sinica，2019）及改善胰岛β细胞功能（European Journal of Medicinal Chemistry，2017）、山荷叶素（diphyllin）阻断线粒体自噬从而促进米色脂肪分化与产热（Frontier in Endocrinology,2020）、杜鹃杂萜（rhodomeroterpene）通过炎症免疫保护肾小管上皮细胞损伤从而改善急性肾功能丢失症与肾纤维化进展（FASEB Journal,Accepted），为中药民族药的药理物质基础鉴定与作用机制研究提供了重要线索。鉴于天然产物改善线粒体质量控制与肝细胞功能之间的重要线索，首次揭示死亡相关凋亡激活蛋白激酶DRAK2通过RNA可变剪接介导线粒体mtDNA复制与生物合成障碍，导致非酒精性脂肪肝炎（NASH）的新病理机制（Cell Metabolism，2021），为代谢综合征的药物研发提供了原始创新药物靶标与可能干预治疗手段。

关键词：代谢综合征；天然产物；线粒体；药物靶标；干预手段

通讯作者：李静雅，E-mail：jyli@simm.ac.cn

灵芝抗病毒感染的机制与疫情防控

林志彬

（北京大学医学部基础医学院药理学系，北京 100191）

摘要：灵芝及其有效成分增强机体非特异性免疫和特异性免疫功能，抑制病毒入侵和在体内增殖、破坏，发挥抗病毒作用。此外，体外抗病毒试验发现，灵芝及其有效成分对流感病毒（influenza virus）、单纯疱疹病毒（herpes simplex virus）、乙型肝炎病毒（HBV）、人类免疫缺陷病毒（HIV）、新城疫病毒（Newcastle disease virus）、登革病毒（Dengue virus）、肠病毒（enterovirus）和柯萨奇病毒（Coxsackie virus）具有抗病毒作用。其抗病毒作用的机制涉及抑制病毒对细胞的吸附或穿透细胞、抑制病毒早期抗原的活化、抑制病毒在细胞内合成所需的一些酶的活性和阻碍病毒DNA或RNA复制等，但对宿主细胞无毒性，与已知抗病毒药联用还有协同作用。

最近有研究发现，灵芝多糖RF3（2 mg·L-1）对体外培养的SARS-Cov-2有显著的抗病毒作用，稀释至1280倍仍有抑制活性，对Vero E6细胞无毒性。与对照组比较，口服灵芝多糖RF3可降低感染SARS-Cov-2病毒的仓鼠肺中病毒载量，但不减轻动物体重。另有研究发现，灵芝多糖作为免疫佐剂可促进接种肠病毒EV-A71灭活疫苗（滴鼻）小鼠的特异性抗体IgG和IgA产生，减轻EV-A71感染小鼠症状，并降低死亡率，具有免疫保护作用。

灵芝抗病毒感染作用及其机制研究为临床防治病毒性疾病提供了理论根据，也为防控新冠疫情提供了潜在的候选药。

关键词：灵芝；增强免疫；抗病毒

通讯作者：林志彬，E-mail：linzb1937@sina.com

桑枝总生物碱治疗糖尿病的研究进展

申竹芳

（中国医学科学院北京协和医学院药物研究所，天然药物活性物质与功能国家重点实验室，中国医学科学院糖尿病研究中心，北京 100050）

摘要：“桑枝总生物碱片”是我国原创降血糖天然药物，于2020年3月获批上市，是目前糖尿病治疗领域唯一的有效组分天然药物。我国拥有完全自主知识产权，并且掌握该品种的关键核心技术，先后获中国发明专利以及美国和日本PCT专利授权。“桑枝总生物碱及其制剂”从立项到上市历经21年，体现了“传承精华，守正创新”的中医药发展理念。桑枝生物总生物碱是以种桑养蚕过程中的废弃物“桑枝”为药材，经提取、分离、纯化而得。在非临床研究中，采用了多种现代药理学和药学研究方法，挖掘出桑枝总生物碱具有作用靶点清晰、物质基础明确、质量可控和降血糖作用显著等特点。明确了其对α葡萄糖苷酶具有极强的抑制活性，且靶点更为精准作用于双糖酶。由北京协和医院牵头完成的多中心临床试验，采用了国际公认的“糖化血红蛋白”金指标，无论单独使用还是用于二甲双胍控制不佳的联合治疗，均具有良好的降糖化血红蛋白效果，且不良反应显著低于对照药物阿卡波糖。此外，单药及联合用药可更及时地改善2型糖尿病患者的餐后1 h血糖，还显示出良好的降低体重、调节糖脂代谢等多种综合获益，突显了天然药物独特的多重药理作用优势。

因此，后续采用了多种现代药理学手段，通过系统性药理学的基础研究，广泛地挖掘桑枝总生物碱的多重药理作用特点与潜在机制。从糖耐量异常、心血管受益、糖脂代谢、肠道菌群、糖尿病并发症等多个层面，深入阐明桑枝总生物碱抗糖尿病作用的物质基础与作用机理。基于现有研究结果推测，桑枝总生物碱片可能直接或间接通过调节营养物质的停留和吸收时间，影响肠道菌群微生态及其代谢产物的变化，进而影响脑-肠-胰岛轴降糖激素的分泌，达到调节脂质吸收和代谢及全身炎症状态等作用；另外，药代动力学研究表明，桑枝总生物碱的主要组分在体内的吸收和多个组织的分布情况，也奠定了其在机体发挥多重药理作用的物质理论基础，最终达到其综合抗糖尿病作用及全身获益。除了对血糖稳态的维持以外，桑枝总生物碱很可能对代谢综合征、糖尿病微血管及大血管并发症等适应症均具有良好调节作用。

桑枝总生物碱片的研究立足中医药，聚焦现代天然药物，桑枝变废为宝，探索绿色循环的中国原创新药发展路径，充分发挥天然药物对复杂疾病治疗的临床优势，具有重要示范引领作用和重大社会效益。因此，前期的研究成果“桑枝总生物碱对糖尿病患者具有降糖化血红蛋白疗效并正式应用于临床”入选了中国2020年度重要医学进展。后续，我们的团队也将继往开来，围绕机制探索及挖掘新适应症等开展研究工作，为促进我国自主知识产权创新中药的上市后再评价及其临床应用，提供更切实的非临床实验数据支持和理论依据。

关键词：桑枝总生物碱；2型糖尿病；天然药物；多重药理作用；综合获益

通讯作者：申竹芳，E-mail：shenzhf@imm.ac.cn

大麻二酚在胶质瘤中的抗肿瘤作用及其机制研究

黄腾飞，闫 超

（南京大学生命科学学院，江苏 南京 210023）

摘要：目的大麻二酚（CBD）是从植物大麻中提取的560余种化合物之一。CBD是一种非精神活性化合物，并且具有很好神经系统保护作用，2018年，CBD被美国FDA批准用于治疗小儿难治性癫痫。由于CBD具有容易通过血脑屏障、安全性高、有一定的抗肿瘤活性等特点，我们认为CBD具有治疗胶质瘤的潜力。本研究对CBD治疗胶质瘤的机制展开系统研究，评估了其对胶质瘤的抑制效果并剖析了其分子机制。方法使用多种胶质瘤细胞系和人源的胶质瘤原代细胞通过体外和体内实验评价CBD对胶质瘤的抗肿瘤效果。在机制研究方面，一方面使用分子生物学和药理学方法探究了CBD直接杀伤胶质瘤细胞的分子机制；另一方面，使用组学方法探究了CBD通过调控巨噬细胞对胶质瘤细胞的吞噬能力间接抑制胶质瘤的作用。结果首先通过多种方法证明了CBD对胶质瘤的抑癌作用，并发现CBD在胶质瘤动物模型中可显著抑制肿瘤生长并延长小鼠生存期。接下来揭示了CBD可通过诱导胶质瘤细胞发生致死性线粒体自噬抑制胶质瘤生长。分析了CBD诱导胶质瘤细胞发生线粒体自噬的分子机制，发现CBD通过激活TRPV4诱导胶质瘤细胞死亡，进而证明CBD诱导胶质瘤细胞死亡是由TRPV4-ATF4-DDIT3-TRIB3-Akt-mTOR信号轴所介导。最后，评估了CBD与替莫唑胺联合用药对于胶质瘤的治疗效果，发现两者联合相较于单药具有更显著的抑癌效果，表现出了较好的临床应用前景。其次，探究了CBD对胶质瘤免疫微环境的影响。肿瘤细胞能够通过躲避巨噬细胞的吞噬作用实现免疫逃逸。体内外研究发现，CBD处理可显著增强巨噬细胞对肿瘤细胞的吞噬作用。在机制方面，发现CBD可通过影响胶质瘤脂代谢稳态导致细胞质膜的稳定性异常，从而促进巨噬细胞对肿瘤细胞的吞噬，发挥抗胶质瘤作用。结论对CBD抗胶质瘤的作用机制进行了系统的研究。一方面，发现CBD可通过诱导致死性线粒体自噬抑制胶质瘤细胞生长；另一方面，发现了CBD通过调节脂质代谢抑制肿瘤免疫逃逸的新机制。CBD可通过这2种机制协同抑制胶质瘤形成与发展，在抗胶质瘤方面是一个有很大临床应用前景的小分子药物。

关键词：大麻二酚；胶质瘤；抗肿瘤

通讯作者：闫 超，E-mail：yanchao@nju.edu.cn

改良型新药：中药大品种培育新方向

杨洪军

（中国中医科学院医学实验中心，北京 100700）

摘要：《中药大品种科技竞争力报告》显示，我国中成药产品科技竞争力普遍比较弱，存在严重的两极分化，市场价值与临床价值、科学价值存在错位，亟待进行以质量、工艺、适应症为核心的大品种培育。

2020年，国家药品监督管理局发布了《中药注册分类及申报资料要求》，中药注册按照中药创新药、中药改良型新药、古代经典名方中药复方制剂、同名同方药等进行分类，前3类均属于中药新药。其中，中药改良型新药指改变已上市中药的给药途径、剂型，且具有临床应用优势和特点，或增加功能主治等的制剂。改良型新药的提出，充分体现了完善全生命周期管理、鼓励中药二次开发的基本导向，并赋予老产品技术研发后的新药属性，成为中药大品种培育的新方向。

作者提出了中药大品种培育的343理论模型。临床价值大、科学价值强、市场价值高三位一体的基本价值取向；将中药大品种立足于四方向的核心内涵，进行产品科技赋能，即承载中医原创理论的代表性品种、凸显中医诊疗优势的代表性品种、催生疾病防治策略变革的代表性品种、融入主流医学诊疗体系的代表性品种；围绕技术提升、药物政策、市场营销3个环节，开展系统化布局与体系化实施。

中成药大品种培育的科技重点包括疗效、质量、机理、理论4个方面。具体而言，疗效得到临床广泛认可，质量控制体系健全，作用机理得到深入阐释，相关中医理论有创新发展。其中，中药复杂作用解析是亟待突破的关键科学问题之一。技术进步促进了化学物质、体内过程、作用机制单一环节的研究，但是，中药物质实体与生命活动的整合调节机制，是中药复杂体系作用模式解析未来面临的关键难点与挑战。“药有个性之专长、方有合群之妙用”，复方是中药应用主要形式，建立在中医学对病机过程的全面判断基础上的多味中药的配伍，作用特征表现为多环节、多靶点的整合调节。这种整合调节的物质基础体内“药动-药效”关联性是如何体现的；多环节、多靶点效应累加模式；药效物质对靶点的直接作用，还是药效物质改变了内环境从而影响了靶点（如受体）的生物性能；也可能是方剂中有效成份以低于它们中某一单体治疗剂量进入人体后，有选择地反复作用于影响某种疾病的主要直接靶点和多个间接靶点，动员机体抗病能力，调节失衡状态，从而达到治疗疾病目的；研究中药“分子网络-药理活性-病证效应”多层次整合作用，阐释中药方剂临床疗效产生的基本机制，将揭示中药作用的基本原理，重新认识中药的科学价值。以上问题的有效解决，将对改良型中药新药研发提供创新技术与方法。

关键词：改良型新药；中药大品种

通讯作者：杨洪军，E-mail：hongjun0420@vip.sina.com

源自针灸临床的靶标发现科学路径——以针刺防治哮喘为例

杨永清1，尹磊淼1，朱维良2，王 宇1，徐玉东1，陈艳焦1

（1.上海中医药大学上海市针灸经络研究所，上海 200030；2.中国科学院上海药物研究所，上海 201203）

摘要：目的如何在传承精华的基础上，实现针灸医学守正创新，既要科学认识针灸，从科学出发研究针灸学问题，更要科学发展针灸，从针灸出发研究医学与生命科学问题。以期从针灸出发，发挥中医药原创优势、推动我国生命科学实现创新突破的伟大目标。方法本文结合自身多年对针灸学与生命科学的思考和长期从事针刺防治哮喘工作的实践，基于针灸治疗有效性的针灸效应物质基础研究，系统建立了源自针灸的靶标发现科学路径，并对未来可期的新药创制进行了展望。结果源自针灸临床的靶标发现科学路径，包括经验传承、临床疗效、效应调节、生物过程、物质基础和靶标发现等。①经验传承：承继邵经明教授“三穴五针”法针刺治疗哮喘临床经验，蒙受的不仅仅是名医的学术思想，更是其品德修为、人格魅力的恩泽。②临床疗效：多年的临床研究证明，“三穴五针”法针刺治疗哮喘不仅具有良好的近期疗效，也有一定的远期效果。该方法针刺治疗哮喘已经成为国家中医药管理局适宜推广技术在全国推广应用。③效应调节：“三穴五针”法针刺治疗哮喘能明显改善患者的呼吸功能及免疫学、甲皱微循环和血液流变学指标，促进Th1/Th2细胞免疫状态趋于平衡。④生物过程：利用组学技术发现针刺抗哮喘效应包括了抗原递呈减弱、免疫抑制调节、平滑肌收缩舒张调控等3类关键生物过程。⑤物质基础：针刺治疗哮喘的物质基础包括KLRC1，CC10，S100A8，S100A11，RAGE，S100A9，MT-2和Dusp1等特异性响应基因和应答蛋白。⑥靶标发现：研究确认MT2的受体是肌动蛋白结合蛋白2（TG2）。获得了称为TSG12的TG2激动剂，TSG12舒张气道平滑肌效应明显优于传统β2激动剂。结论上述源自针灸的靶标发现之科学路径，是“传承精华，守正创新”的生动实践和具体事例，不仅在传承的基础上，阐明了针灸科学机制，同时为解决中国生物医药长期缺乏自主知识产权药物靶标的“卡脖子”问题，提供了针灸解决方案。

关键词：针灸；靶标发现；哮喘

通讯作者：杨永清，E-mail：yyq@shutcm.edu.cn

中药复方制剂的注册管理与高质量转化

赵军宁，戴 瑛，华 桦，曾 瑾

（四川省中医药科学院转化药理与临床应用研究所，四川省中医药转化医学中心，国家中医药管理局中药质量生物评价重点研究室，中医药转化医学四川省重点实验室，四川省道地药材形成原理与品质评价工程研究中心，四川省道地药材系统开发工程技术研究中心，四川 成都 610041）

摘要：转化医学旨在提高总体医疗水平，保障患者的健康以及促进基础研究的深入发展。美国及英国、德国等国家均将“推动实验发现用于临床治疗”作为国家医学研究的重要战略，且通过各种手段加大对转化医学的支持和发展的力度。中医药在数千年的医学实践中不断吸收和融合各个时期的先进科学技术和人文思想，不断创新发展，其理论体系日趋完善，技术方法更加丰富，形成了独特的生命观、健康观、疾病观、防治观。从宏观、系统、整体角度揭示了人的健康与疾病的发生发展规律。随着人类疾病谱的变化，传统中医药理法方药及其主要治病手段——中药复方以其在慢性复杂多因素性疾病的治疗方面具有显著优势而备受关注，日益得到包括国际主流医药界的重视和认可。转化医学强调“环境-社会-心理-工程-生物”的整体医学观及复杂理论系统研究的重要性，与几千年来“临床实践→理论认识→临床实践”的传统中医药发展模式不谋而合。传统中医药从神农尝百草、个体化辨证处方到渐次演变、转化而来的现代复方中成药，从五行生克制化理论、运气学说以及经络流注理论的演变，到金元医家的创新，再到温病学说的形成与发展，无不体现着中医学理论来源于临床实践又在临床实践中得到验证和完善的特点，临床与基础的相互转化一直是中医药发展的主线。随着生命组学尤其是免疫组学理论技术的加速度发展，以临床为导向的中医药现代化研究发展迎来了最佳的时代契机，中医整体观与现代医学融合的新型转化医学模式已经开启，这将为具有临床价值中药新药的加速研制提供强大助力。

关键词：中药复方制剂；新药注册管理；转化医学；新药创制

通讯作者：赵军宁，E-mail：zarmy@189.cn

若干中药化学成分与药理活性研究

赵勤实

（中国科学院昆明植物研究所，云南 昆明 650201）

摘要：药典收录中药品种是中药中具有重要研究价值的品种，但药典品种化学成分与药理活性研究仍然不充分。本课题结合天然药物化学、分子药理学和结构药理学，通过化学成分分离、活性筛选、结构修饰、化学生物学方法研究，开展了丹参、土荆皮、泽泻和川芎等若干药典收录品种化学成分与药理活性研究，发现丹参酮、土荆皮甲酸、泽泻醇和苯酞类化合物新的作用机制和靶点，通过结构修饰和构效关系研究，探讨了构效关系，为找到候选药物分子奠定研究基础。

关键词：中药；化学成分；作用机制；构效关系

通讯作者：赵勤实，E-mail：qinshizhao@mail.kib.ac.cn

青年论坛

Forsythiae Fructus inhibits melanoma growth through activating MAPKs/Nrf2/HO-1 signaling and modulating glycerophospholipid metabolism

BAO Jiao-lin1,2,HE Cheng-wei2

（1.School of Pharmaceutical Sciences,Hainan University,Haikou 570228,China；2.State Key Laboratory of Quality Research in Chinese Medicine,Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVEForsythiae Fructus（Lianqiao）is a typical heat-clearing and detoxicating traditional Chinese medicine（TCM）herb,which has been traditionally used for treating cancer according to TCM theory.However,the under⁃lying mechanism has not been fully explained.METHODSIn this study,we investigated the antitumor effect of Forsyth⁃iae Fructus aqueous extract（FAE）on B16-F10 melanoma.RESULTSFAE strongly inhibited the tumor growth and metastasis formation in B16-F10 melanoma transplanted mice.The survival time of tumor-bearing mice was also signifi⁃cantly prolonged by FAE.The levels of ROS,MDA,TNF-α and IL-6 decreased,while GSH increased in the FAE treat⁃ment group,indicating FAE possesses strong anti-oxidative and anti-inflammatory activity.Western blotting analysis demonstrated that antioxidant proteins Nrf2 and HO-1,tumor suppressors P53 and p-PTEN,and the MAPK pathways in tumor tissues were upregulated by FAE treatment.Serum metabolomics analysis further uncovered that 17 metabolites mostly involving in glycerophospholipid metabolism were correlated with the antitumor effect of FAE.Notably,several lysophosphatidylcholines（LysoPCs）significantly decreased in tumor model group,while FAE treatment restored the changes of these phospholipids to about normal condition.LysoPC acyltransferase 1（LPCAT1）and autotaxin（ATX）highly expressed in melanoma and markedly downregulated by FAE were believed to be responsible for this modulation.CONCLUSIONFAE exhibites strong antitumor activity against B16-F10 melanoma through activating MAPKs/Nrf2/HO-1 mediated anti-oxidation and anti-inflammation and modulating glycerophospholipid metabolism via downregulating LPCAT1 and ATX.Besides,it is suggested that serum LysoPCs could be potential biomarkers for the diagnosis and prog⁃nosis of melanoma.

Key words：Forsythiae Fructus;melanoma;anti-oxidation;anti-inflammation;glycerophospholipid metabolism

Corresponding author：HE Cheng-wei，E-mail：chengweihe@um.edu.mo；BAO Jiao-lin,E-mail：baojiaolin@haina⁃nu.edu.cn

Liujunzi decoction ameliorats cisplatin-induced anorexia via adjusting metabolic disorders in rats

DAI Yong-zhao,CHEN Si-qi,NIE Ke

（School of Chinese Materia Medica,Guangdong Pharmaceutical University,Guangzhou 510006,China）

Abstrect：OBJECTIVECisplatin is a formidable chemotherapy agent widely applying in antineoplastic treatments,but its side effects often limit the clinical usage.Metabolic disorders are one of the side effects induced by cisplatin,which closely relate to the onset of chemotherapy-induced anorexia（CIA）in cancer patients but lacks effective controls.Liujunzi decoction（LJZD）is a traditional Chinese formula that has a promising effect in treating CIA.However,whether LJZD ameliorates CIA through adjusting cisplatin-induced metabolic disorders remain unknow.The present study evalu⁃ated the mechanism of cisplatin-induced metabolic disorders,and the effect of LJZD in ameliorating these disturbances.METHODS42 male Sprague-Dawley（SD）rats（180-220 g）were randomly divided into 3 groups：normal control group（distilled water+saline）,model group（distilled water+cisplatin）,LJZD group（4.8 g·kg-1Liujunzi decoction ingredients+cisplatin）.Intragastrical administered each drug twice a day（7∶00-19∶00）since day 0 for 4 d,animals were intraperito⁃neal injected with cisplatin 6 mg·kg-11 h after administration while normal control groups were injected with same volume of saline.On day 3,each group was anesthetized with pentobarbital sodium 45 mg·kg-1（ip）,and blood samples were collected from aorta abdominalis.Then the samples were analyzed using an LC-ESI-MS/MS system.Significantly regu⁃lated metabolites between groups were determined by VIP ≥1 and absolute Log2FC（fold change）≥1.Identified metabo⁃lites were mapped to Kyoto Encyclopedia of Genes and Genomes（KEGG）pathway database using Metaboanalyst 5.0（https：//www.metaboanalyst.ca/）.RESULTSA total of 133,77 and 32 differential metabolites were filtrated in control vs model,control vs LJZD and model vs LJZD groups respectively.Comparing to control,the levels of hexadecanoic acid（Log2FC=6.3153）,linoleic acid（Log2FC=5.3478）,and 8,11-icosadienoic acid（Log2FC=5.2342）significantly increased,and the levels of N-acetyl-L-tyrosine（Log2FC=-2.6283）,cinnamic acid（Log2FC=-2.3381）,N-acetylphenylalanine（Log2FC=-2.2501）significantly decreased in model group.The KEGG pathway enrichments of these metabolites indi⁃cated that,cisplatin-induced metabolic disorders by disturbing metabolism pathways such as linoleic acid metabolism,biosynthesis of unsaturated fatty acids,and phenylalanine metabolism,which suggested that the onset of CIA was partly associated with the metabolic disorders of linoleic acid,unsaturated fatty acids,and phenylalanine.Compared to control,treatment of LJZD significantly increased the levels of 4-hydroxytryptamine（Log2FC=12.0186）,hexadecanoic acid（Log2FC=5.7412）,linoleic acid（Log2FC=5.1877）and significantly decreased the levels of N-acetylmethionine（Log2FC=-1.7317）,2-aminoethanesulfinic acid（Log2FC=-1.6578）,N-acetyl-L-tyrosine（Log2FC=-1.5355）.And com⁃paring to the model group,4-hydroxytryptamine（Log2FC=12.0186）,7,12-diketocholic acid（Log2FC=2.0998）,N-acetylneuraminic acid（Log2FC=2.0560）markedly increased,and 3-hydroxy-3-methylpentane-1（Log2FC=-1.9202）,5-dioic acid（Log2FC=-1.7166）,N-isovaleroylglycine,hexanoyl glycine（Log2FC=-1.4958）markedly decreased in LJZD group.It was worth noting that,there were 23 differential metabolites filtrated both in control vs model and model vs LJZD groups,which were the key metabolites of LJZD in treating CIA.Among these 23 common metabolites,there were 16 metabolites excluding the control vs LJZD group,that was,LJZD had no effect in normal rats while being able to ameliorated cisplatin-induced metabolic disorders by regulating these 16 metabolites.Cisplatin-induced downregula⁃tion of 11 metabolites such as hydrocinnamic acid,（±）12（13）epoxy-9Z-octadecenoic acid,cinnamic acid were upregulated after LJZD treatment,and cisplatin-induced upregulation of imidazoleacetic acid,2′-deoxycytidine-5′-monophosphate and other 5 metabolites were downregulated by LJZD.The KEGG pathway analysis indicated that the linoleic acid metabolism,histidine metabolism,and pyrimidine metabolism were the most enriched metabolic pathway.Thus,cisplatininduced metabolic disturbances mainly by disturbing linoleic acid metabolism,histidine metabolism,and pyrimidine metabolism,and LJZD interacted with these metabolic pathways to reduce metabolic disorders and thus ameliorated CIA.CONCLUSIONCisplatin-induced anorexia was closely related to the metabolic disorders of linoleic acid metabo⁃lism,biosynthesis of unsaturated fatty acids,and phenylalanine metabolism.The mechanism of LJZD in ameliorating CIA was in concerned with the metabolic adjustments,relating to the regulation of linoleic acid metabolism,histidine metabolism,and pyrimidine metabolism.

Key words：Liujunzi decoction;chemotherapy-induced anorexia;cisplatin;metabolomic;metabolic disorders

Foundation item：National Natural Science Foundation of China（82174143）;and Scientific Research Foundation of Guangdong Pharmaceutical University（51348136）

Corresponding author：NIE Ke,E-mail：nicknk@hotmail.com

Osthole attenuates pulmonary arterial hypertension by modulation of phospholipid metabolism

FU Min,LI Yuan,YAO Li

（Department of Medicinal Chemistry and Natural Medicine Chemistry,Department of Pharmacognosy,College of Pharmacy,Harbin Medical University,Harbin 150081,China）

Abstract：OBJECTIVEPulmonary arterial hypertension（PAH）is a malignant pulmonary vascular disease lacking efficacy therapeutics.Therefore,it urgently needs to develop safe and effective drugs for PAH treatment.Osthole derived from Cnidium monnieri（L.）Cusson（Shechuangzi）or Angelica pubescens Maxim（Duhuo）has the capacity to alleviate PAH by decreasing pulmonary arterial pressure and alleviating pulmonary vascular remodeling in rats,which is a candi⁃date drug for the prevention of PAH,but the underlying modulatory mechanism is still unclear.Our study aims at investi⁃gating the metabolic modulatory mechanism of osthole against PAH employing functional metabolomics strategy.METH⁃ODSPAH model rats were successfully established with MCT,following osthole administration,then functional metabo⁃lomics based on untargeted metabolomics assay,targeted lipidomics analysis,qRT-PCR,Western blotting and ELISA were performed to investigate the modulatory mechanism of osthole against pulmonary arterial pressure and pulmonary vascular remodeling in PAH.RESULTSUntargeted metabolomics results found that sphingosine 1-phosphate（S1P）was the differential metabolites characterized PAH and reversed by osthole treatment.S1P is a crucial sphingolipid metabolite catalyzed by sphingosine kinases1（Sphk1）and functions as promoting PASMCs proliferation contributing to pulmonary vascular remodeling and pulmonary arterial pressure increase.We revealed that osthole reversed high level of S1P by modulating metabolic enzyme Sphk1 via inactivating microRNA-21-PI3K/Akt/mTOR signal pathway to decrease pulmonary arterial pressure in rats with PAH.Then,targeted phospholipid metabolomics results uncovered that decadienyl-L-carnitine（C10：2）was the differential metabolite characterized PAH and corrected by osthole treatment in rat with PAH.C10：2 is the intermediate metabolite of fatty acid oxidation（FAO）,and C10：2 accumulation indicated mitochondrial dysfunction and FAO increase.CONCLUSIONOsthole could block lipid metabolic reprogramming through functional modulating the expression of fatty acid translocase,fatty acid synthase,phospholipase A2,carnitine palmitoyltransferase 1A to inhibit C10：2,thus to improve mitochondrial dysfunction and inhibit utilizing lipid to biosyn⁃thesize necessary essence for pulmonary artery smooth muscle cells（PASMCs）proliferation.Moreover,we delineated that C10：2 and metabolic reprogramming enzymes were modulated by miRNA-22-3p which was involved in PASMCs proliferation and pulmonary vascular remodeling.Therefore,osthole inhibited miRNA-22-3p mediated lipid metabolic reprogramming to ameliorate pulmonary vascular remodeling.

Key words：osthole;pulmonary arterial hypertension;functional metabolomics;phospholipid metabolism

Corresponding author：YAO Li,E-mail：liyao.prof@hotmail.com

Compound Kushen injection induces immediate hypersensitivity reaction through promoting the production of platelet-activating factor via de novo pathway

GAO Yuan1＊,HAI Li-na2,3＊,KANG Yuan1,QIN Wen-jie3,LIU Fang3,CAI Run-lan1,YANG Xiu-wei2,4,QI Yun1

（1.Institute of Medicinal Plant Development,Chinese Academy of Medical Sciences & Peking Union Medical College,Beijing 100193,China;2.Academy for Advanced Interdisciplinary Studies,Peking University,Beijing 100871,China;3.Beijing Zhendong Guangming Pharmaceutical Research Institute,Beijing 100085,China;4.State Key Laboratory of Natural and Biomimetic Drugs,Department of Natural Medicines,School of Pharmaceutical Sciences,Peking University,Beijing 100191,China）

Abstract：OBJECTIVECompound Kushen injection（CKI）is a bis-herbal formulation extracted from Kushen（Radix Sophorae Flavescentis）and Baituling（Rhizoma Heterosmilacis Japonicae）.Clinically,it is used as the adjuvant treat⁃ment of cancer.However,with the increased application,the cases of immediate hypersensitivity reactions（IHRs）also gradually rise.In this study,we investigated the underlying mechanism（s）and active constituent（s）for CKI-induced IHRs in experimental models.METHODST helper 2（Th2）immunity-amplified mice were prepared by aluminum adjuvant.Anaphylactic shock was detected by measuring rectal thermometry in propranolol pretreated mice.For evaluating micro⁃vascular permeability,Evans blue extravasation assay was used.Platelet-activating factor（PAF）,serum total IgE（tIgE）and mouse mast cell protease 1（MMCP1）were measured by ELISA.RESULTSThe obtained results showed that CKI did not elevate serum tIgE and MMCP1 after consecutive immunization for five weeks,but could induce Evans blue extravasation（local）and cause obvious hypothermia（systemic）after a single injection.Further study showed that alka⁃loids in Kushen,especially matrine,were responsible for CKI-induced IHRs.Mechanism study showed that various PAF receptor antagonists could significantly counter CKI-induced IHRs locally or systemically.In cell system,CKI was able to promote PAF production in a non-cell-selective manner.In cell lysate,the effect of CKI on PAF production became stron⁃ger and could be abolished by blocking de novo pathway.CONCLUSIONIn conclusion,our study identifies,for the first time,that CKI is a PAF inducer.It causes non-immunologic IHRs,rather than IgE-dependent IHRs,by promoting PAF production through de novo pathway.Alkaloids in Kushen,especially matrine,are the prime culprits for IHRs.Our find⁃ings may provide a potential approach for preventing and treating CKI-induced IHRs.

Key words：Compound Kushen injection;matrine;platelet-activating factor;non-immunologic immediate hypersen⁃sitivity reactions;de novo pathway

＊Co-first author.

Desulfovibrio vulgaris,a potent acetic acid-producing bacterium,increased by Astragalus polysaccharides to attenuate nonalcoholic fatty liver disease in mice

HONG Ying1,SHENG Li-li1,ZHENG Ning-ning1,JIA Wei2,3,LI Hou-kai1

（1.Functional Metabolomic and Gut Microbiome Laboratory,Institute of Interdisciplinary Integrative Medicine Research,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China;2.Shanghai Key Laboratory of Diabetes Mellitus and Center for Translational Medicine,Shanghai Jiao Tong University Affiliated Sixth People′s Hospital,Shanghai 200233,China;3.School of Chinese Medicine,Hong Kong Baptist University,Hong Kong 999077,China）

Abstract：OBJECTIVEAlthough the underlying mechanism is largely unknown,gut dysbiosis has emerged as a central initiator of obesity-related diseases including nonalcoholic fatty liver disease（NAFLD）,type 2 diabetes and meta⁃bolic syndrome.The emerging evidence support the use of prebiotics like herb-derived polysaccharides for treating NAFLD by modulating gut microbiome.So,our study focused on the microbiota-dependent anti-NAFLD effect and the exact mechanisms of Astragalus polysaccharides（APS）extracted from Astragalus mongholicus Bunge in high-fat diet（HFD）fed mice.METHODSCo-housing experiment was used to assess the microbiota dependent anti-NAFLD effect of APS.Then,targeted metabolomics and metagenomics were adopted for determining short-chain fatty acids（SCFAs）and bacteria that were specifically enriched by APS.Further in vitro experiment was carried out to test the capacity of SCFAs-producing of identified bacterium.Finally,the anti-NAFLD efficacy of identified bacterium was tested in HFD fed mice.RESULTSOur results first demonstrated the anti-NAFLD effect of APS in HFD fed mice and the contribution of gut microbiota.Moreover,our results indicated that SCFAs,predominantly acetic acid were elevated in APS-supplemented mice and ex vivo experiment.Metagenomics revealed that D.vulgaris from Desulfovibrio genus was not only enriched by APS,but also a potent generator of acetic acid,which showed significant anti-NAFLD effects in HFD fed mice.In addition,D.vulgaris modulated the hepatic gene expression pattern of lipids metabolism,particularly suppressed hepatic fatty acid synthase（FASN）and CD36 protein expression.CONCLUSIONAPS enriched D.vulgaris is effective on attenuating hepatic steatosis possibly through producing acetic acid,and modulation on hepatic lipids metabolism in mice.Further studies are warranted to explore the long-term impacts of D.vulgaris on host metabolism and the underly⁃ing mechanism.

Key words：Desulfovibrio vulgaris;Astragalus polysaccharides;nonalcoholic fatty liver disease;

Fundation item：National Natural Science Foundation of China（81873059;82004016）

Corresponding author：ZHENG Ning-ning,E-mail：zhengning201109@163.com;JIA Wei,E-mail：weijia1@hkbu.edu.hk;LI Hou-kai,E-mail：hk_li@shutcm.edu.cn

Chaetocin：a review of its anticancer potential and mechanisms

JIANG Hang-yu1,2,LI Yu-qi1,2,XIANG Xiao-cong3,TANG Zhi-li1,LIU Kang3,SU Qiang1,ZHANG Xiao-fen4,LI Lin1,5

（1.Department of Pharmacy,3.Institute of Tissue Engineering and Stem Cells,4.Department of Urology,The Second Clinical Medical College of North Sichuan Medical College,Nanchong 637000,China;2.School of Pharmacy,North Sichuan Medical College,Nanchong 637100,China;5.College of Bioengineering,Chongqing University,Chongqing 400016,China）

Abstract：Chaetocin is a natural metabolite product with various biological activities and pharmacological functions isolated from Chaetomium species fungi belonging to the thiodiketopyrazines.Numerous studies have demonstrated a wide range of antitumor activities of chaetocin in vitro and in vivo.Several studies have demonstrated that chaetocin sup⁃presses the growth and proliferation of various tumour cells by regulating multiple signalling pathways related to tumour initiation and progression,inducing cancer cell apoptosis（intrinsic and extrinsic）,enhancing autophagy,inducing cell cycle arrest,as well as inhibiting tumour angiogenesis,invasion and migration.The antitumor effects and molecular mechanisms of chaetocin are reviewed and analysed in this paper,and the prospective applications of chaetocin in cancer prevention and therapy are also discussed.Our review provides the theoretical basis for exploiting the clinical applica⁃tion of chaetocin in cancer treatment.

Key words：chaetocin;antitumor;apoptosis;signalling pathway

Corresponding author：LI Lin，E-mail：Linli82@cqu.edu.cn;ZHANG Xiao-fen，E-mail：spacewalker@tom.com

Herba Epimedii′s biotransformation in vitro simulated gastrointestinal digestion and faecal fermentation systems

LI Ben1,ZHANG Xiao-yao1,Alberto CP DIAS1,2,ZHANG Xiao-ying1,2,3

（1.Chinese-German Joint Laboratory for Natural Product Research,College of Biological Science and Engineering,Shaanxi University of Technology,Hanzhong 723000,China;2.Centre of Molecular and Environmental Biology,Department of Biology,University of Minho,Braga 4710-057,Portugal;3.Department of Biomedical Sciences,Ontario Veterinary College,University of Guelph,Guelph,Canada）

Abstrect：OBJECTIVEEpimedium is rich in a variety of beneficial active ingredients,and has been widely used in the ethnopharmacological practices,however,its biotransformation in gastrointestinal digestions remain unclear.This study aimed to investigate the dynamic changes of components and biological activity of Epimedium in the in vitro simu⁃lated digestion and subsequent human faecal fermentation.METHODSThe models of in vitro simulated saliva,gastric and intestinal digestion,as well as colonic fermentation were constructed to simulate the digestion process of Epimedium.The dynamic changes of components of Epimedium during the simulated digestions in vitro and subsequent human faecal fermentation were investigated by UPLC-MS,HPLC-DAD combined with principal component analysis（PCA）and multi-ingredient quantitative analysis.RESULTSA variety of metabolites with high contents were produced after 0.5 h of intestinal digestion and colonic fermentation 0.5 h.Application of PCA to HPLC data showed the obvious separation of colonic fermentation 0.5 h stage samples from other colonic fermentation stages samples（24,48 and 72 h）.Addition⁃ally,non-digestion and saliva digestion stage samples clustered together,and there was obvious separation between intestinal digestion samples and gastric digestion samples.The contents of epimedium C,icariin and baohuside I all increased significantly after intestinal digestion[58.70±7.08,47.15±5.68 and（12.78±0.55）mg·g-1]compared with gastric digestion[29.00±5.65,17.40±4.55 and（2.77±0.19）mg·g-1].There were significant differences between sample after 0.5 h of colonic fermentation[64.22±9.32,51.26±6.33 and（16.68±3.19）mg·g-1]and other time points（24,48 and 72 h）in components and the contents of active ingredient,and the content of these components all decreased with the fermentation time.The ability of scavenging ABTS free radicals[IC50=（0.29±0.02）g·L-1]increased significantly compared with gastric digestion[（1.57 ± 0.02）g·L-1],and after 0.5 h of colonic fermentation,the ability also increased significantly.CONCLUSIONGastrointestinal digestion had a significant impact on the contents of active components in Epimedium,and the metabolism of these components mainly occurred in the colon.The intestinal digestion and colonic fermentation significantly improved the anti-ABTS activity of epimedium.

Key words：Epimedium;simulated gastrointestinal digestion;faecal fermentation;biotransformation

Foundation item：Incubation Project on State Key Laboratory of Biological Resources and Ecological Environment of Qinba Areas,China（SLGPT2019KF04-04）;and the ERDF through the COMPETE2020-Programa Operacional Competitividade e Internacionalização（POCI）,Portugal

Corresponding author：ZHANG Xiao-ying,E-mail：zhxying@snut.edu.cn

Anti-tumor activity of Sanguisorba officinalis L.in non-small cell lung cancer and induced apoptosis via PI3K/Akt/mTOR signaling pathway

LI Hong1,LI Jing1,WU Jian-ming1,2

（1.School of Pharmacy,Southwest Medical University,Luzhou 646000,China;2.The Key Laboratory of Medical Electrophysiology,Medical Key Laboratory for Drug Discovery and Druggability Evaluation of Sichuan Province,Luzhou Key Laboratory of Activity Screening and Druggability Evaluation for Chinese Materia Medica,Ministry of Education of China,Institute of Cardiovascular Research,Luzhou 646000,China）

Abstract：OBJECTIVETo investigate the pharmacological effect and mechanism of Sanguisorba officinalis L.（SOL）in non-small cell lung cancer（NSCLC）in vitro and in vivo based on network pharmacology.METHODSNetwork pharmacology was used to analyze the improving effect of SOL on NSCLC and possible targets.Cell counting kit 8（CCK-8）and 5-ethynyl-2′-deoxyuridine（EdU）staining,Western blotting,flow cytometry of Annexin Ⅴ/PI,Hoechst 33342/PI staining detection and immunofluorescence were utilized in vitro.H&E staining,immunohistochemistry staining and Western blotting were performed in vivo.RESULTSBased on network prediction,we analyzed the 208 common targets of SOL and NSCLC.36 core targets in 208 common targets were obtained through cytoscape analysis.And the top 10 core targets included Akt,mTOR,EGFR,etc..KEGG analysis showed that PI3K-Akt signaling pathway was the most likely pathway.Furthermore,the mechanism study found that SOL could activate the PI3K/Akt/mTOR signaling pathway in vitro and in vivo.The anti-proliferative effect of SOL in A549 and H1299 cells was measured and validated by CCK-8 and EdU assay.Immunohistochemical results of Ki67 showed that SOL effectively inhibited tumor growth in vivo.SOL also significantly inhibited the migration and invasion of A549 and H1299 cells.SOL significantly increased the percentage of cells with PI signal in A549 and H1299,and the process of cell death of A549 cells indicated that SOL induced apoptosis.The PARP-1 and caspase-3 in A549 and H1299 were found to be activated in a dose manner.The results in vivo were consistent with those in vitro.CONCLUSIONSOL-induced,caspase-3-mediated apoptosis via the induction of the PI3K/Akt/mTOR signaling pathway in NSCLC,which further clarified the mechanism of SOL in the inhibition of NSCLC,and thereby provided a possibility for SOL to serve as a novel therapeutic agent for NSCLC in the future.

Key words：Sanguisorba officinalis L.;non-small cell lung cancer;network pharmacology;PARP-1;PI3K/Akt/mTOR

Fundation item：National Natural Science Foundation of China（81774013;81804221;82074129）;and National Science and Technology Major Project of China（2018ZX09721004-006-004）

Corresponding author：WU Jian-ming，E-mail：jianmingwu@swmu.edu.cn

In vitro and in vivo anti-HMPV activity of JH001 isolated from Chinese medicinal herbs

LI Jin-hua1,ZHAO Yao4,DAI Ying2,3,ZHAO Jun-ning1,2,3

（1.West China School of Pharmacy Sichuan University,Chengdu610041,China;2.Sichuan Academy of Chinese Medicine Sciences,Chengdu 610041,China;3.Sichuan Institute for Translational Chinese Medicine,Chengdu 610041,China;4.Children′s Hospital of Chongqing Medical University,Chongqing 400014,China）

Abstract：OBJECTIVEHuman metapneumovirus（hMPV）is semblable to respiratory syncytial virus（RSV）which causes respiratory infections typically characterized by cough,runny nose,fever,and nasal congestion but sometimes progressing to bronchiolitis and pneumonia.Whereas,there is no corresponding drug to inhabit the virus.Studies of new compounds with potential anti-HMPV activity could produce clinical value.Chinese herbal medicine played a great role during COVID-19,therefore we choose some small molecular（JH001）extracted from botany to investigate therapeutic effect on hMPV and the underlying mechanisms.METHODSIn this study,16HBE cells were used as a model to explore in vitro antiviral effect.Cytotoxicity assays were performed before the antiviral tests,cell viability of 16HBE cells handled by different concentration of JH001 was estimated by Cell Counting Kit-8（CCK-8）.Then RT-qPCR,immunofluores⁃cence,and flow cytometer were used to test the viral titer after cells infected with hMPV.Eventually,6-8 weeks mice were infected intranasally with 60 μL of hMPV,the control group was treated with 0.9%saline water,other groups were administered with JH001 and ribavirin,then the lung virus titer and protective effect in lung were judged.RESULTSThe obtained JH001 exhibited no cytotoxicity to 16HBE cells during 6.25-200 μmol· L-1.RT-QPCR demonstrated that JH001 showed obvious inhabitation to the viral replication and showed great significance compared with saline.And fluo⁃rescence exhibited distinct decrease of hMPV-N protein,flow cytometer results showed that MFI decrease evidently.Sig⁃nificant reduction of N-gene expression was observed in those mice treated with JH001 compared with saline group,which indicated that JH001 probably had protective and therapeutic effect on viral replication.CONCLUSIONThis study illustrated that JH001 might be a promising option for small molecular against hMPV and JH001 might be worthy of fur⁃ther development and used as a potential therapeutic strategy for other respiratory viruses in the future.

Key words：human metapneumovirus;Chinese herbal medicine;antiviral activity;respiratory infections

Corresponding author：ZHAO Yao,E-mail：nhhco@126.com;DAI Ying,E-mail：837498533@qq.com;ZHAO Jun-ning,E-mail：zarmy@189.cn

Serum proteomic approach in patients with Qi deficiency and blood stasis syndrome in coronary heart disease：to explore therapeutic mechanism of Yiqi Huoxue decoction

LI Ying1,WANG Zhi-bo1,ZHU Ming-jun2,WANG Yong-xia2,MA Bo1,MIAO Lan1,PAN Ying-hong3,LIU Jian-xun1

（1.Beijing Key Laboratory of Pharmacology of Chinese Materia Region,National Clinical Research Center for Chinese Medicine Cardiology,Xiyuan Hospital,China Academy of Chinese Medical Sciences.Beijing 100091,China;2.The First Affiliated Hospital of Henan University of Chinese Medicine,Zhengzhou 450011,China;3.Institute of Crop Science,Chinese Academy of Agricultural Sciences,Beijing 100081,China）

Abstract：OBJECTIVETo explore the curative effect and mechanism of Yiqi Huoxue decoction in the treatment of coronary heart disease with Qi deficiency and blood stasis syndrome.METHODSThe patients with coronary heart dis⁃ease of Qi deficiency and blood stasis syndrome were treated with Yiqi Huoxue decoction for 3 months,and the changes of cardiac function were observed.61 serum samples（including 29 cases of disease group and 32 cases of Yiqi Huoxue expression group）were analyzed by non labeled proteomics.The disease group was used as the control group,and the protein with expression level difference of more than 1.2 folds（P＜0.05）was screened.The molecular function,biologi⁃cal pathway and protein interaction of the different proteins were analyzed by bioinformatics,so as to identify the molecu⁃lar and biological pathway of Yiqi Huoxue decoction in the treatment of coronary heart disease with Qi deficiency and blood stasis syndrome.RESULTSClinical treatment found that Yiqi Huoxue decoction can improve TCM syndrome score and left ventricular ejection fraction,regulate blood glucose and blood lipid levels,prolong thrombin time,and improve heart function.The results of proteomic quantitative analysis showed that there were 69 proteins with different expression levels in the disease group.Bioinformatics analysis results showed that Yiqi Huoxue decoction may regulate ApoA1,alpha-2 and other proteins to act on HDL assembly,platelet degradation,PI3K Akt signaling pathway,and then play a therapeutic role in coronary heart disease with Qi deficiency and blood stasis syndrome.CONCLUSIONYiqi Huoxue decoction can effectively improved the heart function decline caused by Qi deficiency and blood stasis syn⁃drome of coronary heart disease.It mainly act on energy metabolism and platelet activation pathway by activating HDL assembly and platelet degradation signal pathway proteins.This study can provide reference for the follow-up treatment mechanism of Qi deficiency and blood stasis syndrome of coronary heart disease.

Key words：Qi deficiency and blood stasis syndrome;coronary heart disease;Yiqi Huoxue decoction;proteomic

Foundation item：National Natural Science Foundation of China（82030124）;and National Key Basic Research Special Foundation of China（2015CB554400）

Corresponding author：PAN Ying-hong,E-mail：panyinghong@caas.cn;LIU Jian-xun,E-mail：liujx0324@sina.com

A metabolite of Danshen formulae（IDHP）induces angiogenesis and protects rat brains against focal ischemia via CaMKK β/AMPK（Thr172）/eNOS（Ser1177）signaling

LIAO Sha1,2,LIU Rui-min1,XU Dan-ni1,ZHU Ming-hui1,ZHAO Qi1,LUO Xian-lin1,LI Zhu1,LUO Quan-li1,FAN Tai-ping2,ZHENG Xiao-hui1

（1.College of Life Sciences,Northwest University,Xi′an 710069,China;2.Angiogenesis and Chinese Medicine Laboratory,Department of Pharmacology,University of Cambridge,Cambridge,UK）

Abstract：OBJECTIVEOnly limited number of drugs are currently available for treating ischemic stroke.Therapeu⁃tic angiogenesis has recently emerged as one of the most promising therapies for cerebral ischemic injury.Isopropyl-β-（3,4-dihydroxyphenyl）-α-hydroxypropanoate（IDHP）is a metabolite derived from the botanical formulation for Dantonic®.Here,we investigated the angiogenic efficacy of IDHP in cerebral ischemia.METHODSThe in vivo effects of IDHP were evaluated in the C57BL/6 mouse Matrigel plug and rat transient middle cerebral artery occlusion（tMCAO）models.Primary human umbilical vein endothelial cells（HUVEC）and human brain microvascular endothelial cells（HBMEC）were used to explore the effects of IDHP on stimulating proliferation,migration and tube formation in vitro.ELISA and Western blotting were used to quantitate the release and expression of relevant target molecules and signaling path⁃ways.RESULTSIDHP reduced infarct volume and improved sensorimotor function in rats subjected to tMCAO by pro⁃moting angiogenesis,and promoted Matrigel neovascularization in mice.Moreover,IDHP produced a biphasic modula⁃tion on proliferation and migration both in HUVEC and HBMEC.It also induced tube formation in a 12-day HUVEC-HDF co-culture model and in Matrigel assays.IDHP-induced angiogenesis was accompanied by increased levels of p-AMPKα（Thr172）and p-eNOS（Ser1177）both in vitro and in vivo,and the decreased level of VEGF in rat brains on day 1 whereas enhanced level of VEGF on day 3 and 7 after tMCAO.Mechanistically,AMPK knockdown or pharmacologi⁃cally inhibiting AMPK and its upstream kinases（CaMKKβ）inhibited the eNOS phosphorylation induced by IDHP in HUVEC.Furthermore,selective eNOS inhibitor（L-NIO）,selective CaMKKβ inhibitor（STO）and AMPKa inhibitor（Com⁃pound C）blocked the capillary-like tube formation in the co-culture model induced by IDHP（10 nmol·L-1）.CONCLU⁃SIONCollectively,these findings showed that IDHP protected rats from cerebral ischemia-reperfusion injury by promot⁃ing angiogenesis via activating CaMKKβ/AMPK（Thr172）/eNOS（Ser1177）signaling,and suggest it to be a promising new drug candidate for the prevention and/or treatment of cerebral ischemia and other vascular occlusive diseases.

Key words：ischemic stroke;angiogenesis;endothelial cells;functional recovery

Foundation item：National Natural Science Foundation of China（31971143）;and Primary R&D Plan of Shaanxi Province（2021KWZ-24）

Corresponding author：ZHENG Xiao-hui,E-mail：zhengxh318@nwu.edu.cn

Genetic polymorphisms of pharmacogenomic very important pharmacogenes variants in Chinese Lisu population

LU Hong-yan1,2,WANG Yu-liang1,2,ZHANG Zhan-hao1,2,XING Shi-shi1,2,LI Dan-dan1,2,WANG Li1,2,JIN Tian-bo1,2

（1.Key Laboratory of Molecular Mechanism and Intervention Research for Plateau Diseases of Tibet Autonomous Region,School of Medicine,Xizang Minzu University,Xianyang 712082,China;2.Engineering Research Center of Tibetan Medicine Detection Technology,Ministry of Education,Xizang Minzu University,Xianyang 712082,China）

Abstract：OBJECTIVEThe specificity of drug therapy in individuals and races has promoted the development and improvement of pharmacogenomics and precision medicine.While there is a few cognition on the minorities in China,especially in Lisu nationality from the Yunnan province.Therefore,we performed the research to improve the role of pharmacogenomics in the Lisu population from the Yunnan province of China.METHODS54 variants of very important pharmacogenes selected from the PharmGKB database were genotyped in 199 unrelated and healthy Lisu adults from the Yunnan province of China,and then,genotyping data wtih χ2test were analyzed.RESULTSWe compared our data with those of other 26 populations from the 1000 Genomes Project,and acquired that the Lisu ethnicity is similar with the CDX（Chinese Dai in Xishuangbanna,China）and CHS（Southern Han Chinese,China）.Furthermore,rs776746（CYP3A5）,rs1805123（KCNH2）,rs4291（ACE）,rs1051298（SLC19A1）and rs1065852（CYP2D6）were deemed as the most varying loci.The MAF of“G”at rs1805123（KCNH2）in the Lisu population was the largest with the value of 51.0%.CONCLUSIONThere are significant differences in single nucleotide polymorphism loci,supplementing the phar⁃macogenomic information of the Lisu population in Yunnan province,China,and can provide a theoretical basis for indi⁃vidualized medication in the future.

Key words：very important pharmacogenes variants;Lisu population;pharmacogenomics;racial difference

Corresponding author：JIN Tian-bo,E-mail：tianbo__jin@163.com

Desmethylbellidifolin protects against chronic alcoholic fatty liver disease by regulating Akt-mTORC1 pathway mediated autophagy

SHEN Jia-yan,YUAN Ruo-lan,LIU Miao,WANG Tao,LIU Meng-yang

（Tianjin State Key Laboratory of Modern Chinese Medicine,Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China）

Abstract：OBJECTIVETo investigate the protective effect and potential mechanism of desmethylbellidifolin（DMB）in chronic alcoholic fatty liver disease.METHODSC57BL/6 mice were randomly divided into five groups.Control,meta⁃doxine and DMB group（high dose and low dose）mice were fed with Lieber-DeCarli liquid diet containing 5%alcohol for six weeks.Pair-fed group mice were fed with a liquid diet containing the same calories.After treatment,serum GOT,GPT,TG and hepatic T-CHO,TG,GSH,GSH-Px,SOD and CAT levels were measured.Ectopic liver lipid deposition was determined by oil red O and hematoxylin-eosin（HE）staining.Lipid metabolism and autophagy related genes expression were determined by real-time PCR and Western blotting.Electron microscope and laser scanning confocal microscope were used to detect autophagosome and autophagy flux.RESULTSDMB treatment markedly reduced serum TG,GOT and GPT levels in alcohol-induced mice,as well as hepatic levels of T-CHO,TG and MDA,while increased the GSH,GSH-Px,SOD and CAT levels in the liver.Oil red O and HE staining showed that the alcoholinduced lipid accumulation and hepatocyte morphology changes were significantly improved by DMB treatment.Mecha⁃nistically,the expression of stearoyl-CoA desaturase 1 and fatty acid synthase were significantly decreased,while lipoly⁃sis related hormone-sensitive lipase was elevated in mouse liver by DMB treatment.In addition,DMB could inhibit the phosphorylation of Akt and mTORC1,and activate autophagy process by inducing autophagy related genes expression,such as LC3,ATG5 and ATG7.Moreover,treatment with DMB notably increased the number of autolysosome and promote the autophagy flux,which may therefore induce the lipolysis and oxidation of lipids and prevent the alcoholinduced excessive lipid accumulation in the liver.CONCLUSIONDMB exerts a protective role in alcoholic fatty liver dis⁃ease by regulating the Akt-mTORC1 pathway mediated autophagy activation.

Key words：desmethylbellidifolin;alcoholic fatty liver;autophagy;lipid metabolism

Corresponding author：LIU Meng-yang,E-mail：liumengyang0212@tjutcm.edu.cn

Paeonol attenuated vascular fibrosis through regulating Treg/Th17 balance in gut microbiota-dependent manner

SHI Xiao-yan1,2,HUANG Han-wen1,2,ZHOU Min1,2,LIU Ya-rong1,2,WU Hong-fei1,2,DAI Min1,2

（1.College of Pharmacy,Anhui University of Chinese Medicine,Hefei 230012,China;2.Anhui Key Laboratory for Research and Development of Traditional Chinese Medicine,Hefei 230012,China）

Abstract：OBJECTIVEAtherosclerosis（AS）is a chronic inflammatory disease characterized by the accumulation of lipids,vascular fibrosis,and inflammation.Paeonol（Pae）is a natural phenolic compounds isolated from a traditional Chinese medicine,Cortex Moutan,which exhibits anti-AS effects.Our previous work demonstrated that gut microbiota plays an important role during AS treatment as it affects the efficacy of Pae.However,the mechanism of Pae in protect⁃ing against vascular fibrosis as related to gut microbiota has yet to be elucidated.To investigate the anti-fibrosis effect of Pae on AS mice and demonstrate the underlying gut microbiota-dependent mechanism.METHODSApoE-/-mice were fed with high-fat-diet（HFD）to replicate the AS model.HE and Masson staining were used to observe the plaque forma⁃tion and collagen deposition.Gut microbiota alteration and short-chain fatty acids（SCFAs）production were analyzed through 16S rRNA sequencing and LC-MS/MS.The frequency of immune cells in spleen were phenotyped by flow cytometry.The mRNA expression of aortic inflammatory cytokines were detected by qRT-PCR.The protein expression of LOX and fibrosis related indicators were examined by Western blotting.RESULTSPae restricted the development of AS and collagen deposition.Notably,the anti-fibrosis effect of Pae was achieved by regulating the gut microbiota.16S rRNA sequencing and LC-MS/MS data indicated that the relative abundance of SCFAs-producing bacteria and SCFAs production was increased.Additionally,Pae administration selectively up-regulated the frequency of regulatory T（Treg）cells as well as down-regulated the ratio of T helper type 17（Th17）cells in the spleen of AS mice,improving the Treg/Th17 balance.In addition,as expected,Pae intervention significantly down-regulate the mRNA expression levels of proinflammatory cytokines IL-1β,IL-6,TNF-α and IL-17 in the aorta tissue,up-regulate the levels of anti-inflammatory factor IL-10,a marker of Treg cells.Finally,Pae′s intervention in the gut microbiota resulted in the restoration of the balance of Treg/Th17,which indirectly down-regulated the protein expression level of LOX and fibrosis-related indicators（MMP-2/9 and collagen Ⅰ/Ⅲ）.CONCLUSIONPae attenuates vascular fibrosis in a gut microbiota-dependent manner.The under⁃lying protective mechanism is associated with the improved Treg/Th17 balance in spleen mediated through the increased microbiota-derived SCFAs production.

Key words：paeonol;atherosclerosis;gut microbiota;Treg/Th17 balance;vascular fibrosis

Foundation item：National Natural Science Foundation of China（81773937）

Corresponding author：DAI Min,E-mail：daiminliao@163.com

Optimization ratio between Coptis chinensis and cinnamo of Jiaotai pill plus benzodiazepines for insomnia in China：a network meta-analysis

SUN Shi-guang1,2,WANG Qun3,CHEN Li-jing1,2

（1.Department of Pharmacy,Second Affiliated Hospital of Shandong University of Traditional Chinese Medicine,Jinan 250001,China;2.Department of Pharmacy,Shandong Provincial Hospital of Integrated Medicine,Jinan 250001,China;3.Department of Vertigo,Jinan Shizhong People′s Hospital,Jinan 250022,China）

Abstract：OBJECTIVETo explore the efficacy of Jiaotai pill（JTW）and JTW modified prescriptions（JTW+）plus benzodiazepines（BDZ）for insomnia in China by systematic review and network meta-analysis,especially optimization ratio（m：n）between Coptis chinensis and Cinnamo.METHODSRandomized controlled trials（RCTs）regarding JTW and JTW++BDZ for insomina were comprehensively searched in CNKI,VIP,Wanfang,and PubMed,Cochrane Library,OVID,Embase databases from their inception to December 2017.The risk of bias was assessed by the Cochrane collab⁃oration′s tool and Jadad scale.Systematic review and network meta-analysis were performed to evaluate the relative risk（RR）plus 95%confidence interval（95%CI）or prediction interval（95%PrI）,and draw surface under the cumulative rank⁃ing curves（SUCRA）by Stata 14.0 and RevMan 5.0 software.RESULTSA total of 13 RCTs（1 RCT was omitted by sen⁃sitivity analysis）with 1040 patients were included.Overall heterogeneity was acceptable,except publication bias.Based on classical meta-analysis,JTW and JTWm：n++BDZ,such as JTW2：1++BDZ,JTW5：1++BDZ,JTW6：1++BDZ,JTW10：1++BDZ,had more efficacy than BDZ.Based on network meta-analysis,JTW and JTWm：n++BDZ,such as JTW2：1++BDZ,JTW6：1++BDZ,JTW10：1++BDZ,had more efficacy than BDZ;meanwhile,SUCRA showed the best rank of JTW/JTW+（BDZ

Key words：Jiaotai pill;Coptis chinensis;cinnamo;ratio;insomnia;randomized controlled trial;systematic review;network meta-analysis

Fundation item：National Science and Technology Major Project of China（SQ2017ZX090301064）;National Natural Science Foundation of China（81302887）;Shandong Science and Technology Development Program（2014GSF119034）;and Shandong Science and Technology Development Program of Traditional Chinese Medicine（2017-115;2019-0222）

Corresponding author：SUN Shi-guang,E-mail：S.G.SUN@hotmail.com

Identify potential treatments of COVID-19 from Tibetan medicine Hippophae Fructus using a network pharmacological approach

WANG Yu-liang1,WANG Yu-he2,LU Hong-yan1,ZHANG Zhan-hao1,JIN Tian-bo1,WANG Li1

（1.Key Laboratory of Molecular Mechanism and Intervention Research for Plateau Diseases of Tibet Autonomous Region,School of Medicine,Xizang Minzu University,Xianyang 712082,China;2.Department of clinical laboratory,The Affiliated Hospital of Xizang Minzu University,Xianyang 712082,China）

Abstract：OBJECTIVESince the coronavirus disease 2019（COVID-19）outbreak in December 2019,the search for a potential treatment for COVID-19 has been a constant focus.Therefore,we identified potential treatments for COVID-19 from Hippophae Fructus,a Tibetan medicine that may act on COVID-19,using a network pharmacology approach.METHODSWe collected the chemical constituents and corresponding targets of Hippophae Fructus from traditional Chinese medicine system pharmacology（TCMSP）.COVID-19 related genes were predicted in pubmed-Gene,OMIM and GeneCards databases.Then,protein-protein interactions（PPIs）of key genes were analyzed by STRING database.Compound-target-diseases network was constructed using Cytoscape software.The potential pathways were deter⁃mined by Gene Ontology（GO）and Kyoto Encyclopedia of Genes and Genomes（KEGG）pathway analyses.Additionally,molecular docking was used to verify the binding effect between the active component and the target.RESULTSA total of 33 components and 192 corresponding targets in Hippophae Fructus were found.50 genes were obtained from the intersection of component targets and disease targets.These genes include IL-6,TNF,MAPK8 and PTGS2,which regu⁃late several pathways associated with COVID-19,involving Hepatitis B,Influenza A,TNF signaling pathway and Tuber⁃culosis.More importantly,high-node compounds such as quercetin and beta-sitosterol can well bind to key targets.CONCLUSIONSome components in Hippophae Fructus can act on COVID-19 related genes and regulate multiple pathways.Perhaps Hippophae Fructus has the effect in treating COVID-19.

Key words：Hippophae Fructus;COVID-19;network pharmacology;Tibetan medicine;molecular docking

Foundation item：Emergency Research Project on COVID-19 at Xizang Minzu University（XZMDYJ03;and XZM⁃DYJ01）

Corresponding author：WANG Li,E-mail：wangli_xzmd361@163.com

Baicalin protects against 17 α-ethinylestradiol-induced cholestasis via Sirt1/HNF-1 α/FXR pathway

YANG Jin-yu1,XIANG Dao-chun1,2,XIANG Dong1,HE Wen-xi1,LIU Ya-nan1,LAN Lu-lu1,LI Guo-dong1,JIANG Chen3,REN Xiu-hua1,LIU Dong1,ZHANG Cheng-liang1

（1.Department of Pharmacy,Tongji Hospital Affiliated to Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430014,China;2.The Central Hospital of Wuhan,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430014,China;3.College of Pharmacy,Jilin University,Changchun 130021,China）

Abstract：OBJECTIVEBaicalin is a major flavonoid component of Scutellaria baicalensis,and has been used in the treatment of liver diseases for many years.However,the role of baicalin in estrogen-induced cholestasis（EIC）remains to be elucidated.This present study explored the protective effect of baicalin against estrogen-induced liver injury and further elucidated the mechanisms involved both in vivo and in vitro.METHODSWe conducted a series of experiments using 17α-ethinylestradiol（EE）induced cholestatic rats and cultured HepG2 cells.Serum,bile,and liver samples were collected for biochemical and histological analyses.Bile acid composition in liver was analyzed by LC-MS/MS.The mechanisms underlying the hepatoprotective of baicalin were investigated by RT-PCR,Western blotting analyses and immunohistochemistry.RESULTSBaicalin showed obvious hepatoprotective effects in EIC rats by reducing serum bio⁃markers and increasing the bile flow rate,as well as by alleviating liver histology and restoring the abnormal composition of hepatic bile acids（BAs）.In addition,baicalin protected against EE induced liver injury by up-regulation of the expres⁃sion of hepatic efflux transporters and down-regulation of hepatic uptake transporters.Furthermore,baicalin increased the expression of hepatic BA synthase（CYP27A1）and metabolic enzymes（Bal,Baat and Sult2a1）in EIC rats.We showed that baicalin significantly inhibited hepatic inflammatory responses in EIC rats through reducing elevated levels of TNF-α,IL-1β,IL-6 and NF-κB.Finally,we confirmed that baicalin maintains BA homeostasis and alleviates inflamma⁃tion through Sirt1/HNF-1α/FXR signaling pathway.CONCLUSIONBaicalin protects against estrogen-induced cholestatic liver injury,and the underlying mechanism involved is related to activation of the Sirt1/HNF-1α/FXR signaling pathway.

Key words：baiclain;cholestasis;17α-ethinylestradiol;Sirt1;HNF-1α;FXR

Corresponding author：ZHANG Cheng-liang,E-mail：clzhang@tjh.tjmu.edu.cn

Protective effect of Renqing Changjue on acute respiratory distress syndrome in rabbits

YAO Meng-qi1,FU Shu-yan1,ZHOU Zi-qiang1,GENG Sang2,SE Li-ma3,SUN Fang-yun1,BAI-MA Luo-bu2

（1.Engineering Research Center of Tibetan Drug Detection Technology,Ministry of Education,Xizang Minzu University,Xianyang 712082,China;2.Tibet University of Tibetan Medicine,Lhasa 850000,China;3.Tibet Ganlu Tibetan Medicine Co.,Ltd.,Lhasa 851414,China）

Abstract：OBJECTIVETo investigate whether Renqing Changjue has a protective effect on acute respiratory dis⁃tress syndrome（ARDS）induced by endotoxin lipopolysaccharide（LPS）in rabbits.METHODSThirty-six healthy male New Zealand white rabbits were randomly divided into six groups：normal control group,model group,dexamethasone group,Renqing Changjue high,middle and low dose group,with six rabbits in each group.LPS was used to replicate the ARDS model after five consecutive days of gavage.Arterial pressure,respiratory rate and anal temperature blood were recorded for arterial blood gas analysis at 0,0.5,1,2 and 4 h,respectively.At the end of the four-hour experiment,rab⁃bits were killed by bloodletting,and the lung tissue was quickly removed to determine the cytokines,SOD,MDA and pathological examination of rabbit lung.RESULTSRenqing Changjue can significantly reduce the pathological changes of lung in ARDS model group.The expression of AQP1 and MPO in rabbit lung was significantly decreased by immuno⁃histochemistry（P＜0.05），reduce the lung wet/dry weight ratio,increase the ratio of PaO2/FiO2,inhibit the release of inflammatory factors and scavenge free radicals and antioxidant effects.CONCLUSIONRenqing Changjue can effec⁃tively protect rabbits with acute respiratory distress syndrome induced by LPS,and may protect the lung by inhibiting the release of cytokines and anti-oxidation.

Key words：acute respiratory distress syndrome;acute lung injury;cytokines;antioxidant;blood gas analysis

Foundation item：2020 Project of Tibetan Medicine Administration of Tibet Autonomous Region（JJKT202004）;and 2020 Reform and Development Special Project（324042000101）

Corresponding author：SUN Fang-yun,E-mail：xzmysfy@163.com

Suppression of AOM/DSS-induced colorectal cancer by scutellarin through downregulation of Wnt signaling pathway activity

ZENG Sha1,ZHAO Hui1,CHEN Li1,SUN Qiang1,REN Shan1,LIU Mao-lun1,YANG Han1,TANG Shun1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVETo investigate the therapeutic effect of scutellarin on colitis-associated cancer（CAC）and its underlying mechanism based on Wnt/β-catenin signaling pathway.METHODSThe mouse model of CAC was estab⁃lished by azomethane oxide（AOM）and sodium dextran sulfate（DSS）,followed by scutellarin treatment,with recording the body weight,diarrhea and hematochezia.After sacrificing the mice,the colorectal length and colorectal tumor were assessed.The levels of pro-inflammatory factors TNF-α and IL-6 in mice′s sera were measured by the enzyme-linked immunosorbent assay（ELISA）.The colorectal lesions were appraised by hematoxylin and eosin（H&E）staining.The β-catenin level in CAC tissues was probed by immunofluorescent analysis.The apoptosis-related genes Bax and Bcl-2,and Wnt signaling pathway-related genes β-catenin,GSK-3β,TCF4,c-Myc and cyclin D1 were detected by real-time quantitative RT-PCR（RT-qPCR）.Finally,Western blotting analysis（WB）was employed to examine the expressions of the apoptosis and Wnt signaling pathway-related proteins.RESULTSScutellarin significantly improved AOM/DSS-caused weight loss,colorectal length shortening,and tumor growth in mice（P＜0.01）.Meanwhile,colorectal lesions could be substantially alleviated by scutellarin.ELISA results showed that the levels of pro-inflammatory factors TNF-α and IL-6 were drastically lessened（P＜0.01）.Scutellarin also sharply inhibited the nuclear translocation of β-catenin,as evidenced by the reduction in the nuclear level of β-catenin protein.In addition,scutellarin attenuated the mRNA expres⁃sion of Wnt signaling pathway-related β-catenin,TCF4,c-Myc and cyclin D1,whereas it heightened GSK-3β mRNA level.These results were consolidated by WB analysis,which indicated that scutellarin could mitigate the protein levels of phospho-GSK-3β,β-catenin,TCF4,c-Myc and cyclin D1,with the increase in GSK-3β protein in CAC tissue.Moreover,scutellarin could induce the apoptosis of CAC,demonstrated by enhanced expression of Bax and diminished expression of Bcl-2 in both mRNA and protein levels.CONCLUSIONScutellarin may ameliorate colitis-associated colorectal cancer by weakening Wnt/β-catenin signaling cascade.

Key words：scutellarin;colorectal cancer;Wnt signaling pathway;AOM/DSS

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;the Excellent Talent Program of Chengdu University of Traditional Chinese Medicine of China（YXRC2019002;ZRYY1917）;and Open Research Fund of the State Key Labora⁃tory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Novel technologies and models for studying pharmacological activities of traditional Chinese medicine for promoting blood circulation and removing blood stasis

ZHAO Yang1,2,WEI Zhong-hong2,YU Su-yun1,2,WANG Ai-yun2,3,CHEN Wen-xing2,3,ZHENG Xiang-jian4,Mathew VADAS5,Jennifer GAMBLE5,LU Yin2,3

（1.Department of Biochemistry and Molecular Biology,School of Medicine & Holistic Integrative Medicine,2.Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica,School of Pharmacy,3.Jiangsu Collaborative Innovation Center of Traditional Chinese Medicine Prevention and Treatment of Tumor,Nanjing University of Chinese Medicine,Nanjing 210023,China;4.Department of Pharmacology,School of Basic Medical Sciences,Tianjin Medical University,Tianjin 300070,China;5.Centre for the Endothelium,Vascular Biology Program,Centenary Institute,The University of Sydney,Sydney 2050,Australia）

Abstract：OBJECTIVEOur previous studies demonstrated that various ingredients from the traditional Chinese medicine（TCM）for promoting blood circulation and removing blood stasis,as exemplified by cryptotanshinone and salvi⁃anolic acid B,exerted striking effects on modulating angiogenesis and vascular permeability,which suggests that they may be effective in treating vascular leak-driven diseases（e.g.tumor,cerebral cavernous malformation and diabetic reti⁃nopathy）.However,the lack of reliable and advanced technologies and models sets up difficult hurdles for better under⁃standing the role of TCM for promoting blood circulation and removing blood stasis.To this end,this study is to outline numerous cutting-edge platforms that can be utilized for exploring the function of TCM for promoting blood circulation and removing blood stasis in vascular leak-driven diseases.METHODSTwo-photon laser scanning fluorescence micros⁃copy was used to observe the interactions between neutrophils and blood vessels in a real-time manner.Dynamic flow system was employed to mimic the in vivo behaviors of neutrophils.RIP1-Tag5 spontaneous pancreatic cancer model was used to study the function of tumor blood vessels.CCM2ECKO（deletion of CCM2 in endothelial cells）mice were employed to establish the cerebral cavernous malformation（CCM）animal model.Micro-computed tomography（micro-CT）was utilized to assess the CCM lesion.Müller cell-knockout mouse model was used to study the progression of dia⁃betic retinopathy.Vascular permeability in this model was assessed by fluorescein angiography.RESULTSThe interac⁃tions between neutrophils and endothelial cells involve a series of complicated processes,including rolling,adhesion,intraluminal crawling and transmigration,which were all monitored in vivo by two-photon laser scanning fluorescence microscopy in a real-time manner.Dynamic flow system was capable of recapitulating the biological behaviors of neutro⁃phils in vitro.Tumor vascular function in particular vascular perfusion could be assessed in the RIP1-Tag5 spontaneous pancreatic cancer model.In terms of CCM studies,specific deletion of CCM2 in endothelial cells resulted in the initiation of CCM lesion.The size and number of CCM lesions could be visualized and quantified by micro-CT.Furthermore,the Müller cell-knockout mouse model was able to precisely reflect the clinical symptoms of diabetic retinopathy.Vascular leak could be monitored at different time points using fluorescein angiography.CONCLUSIONAn array of high technol⁃ogies and animal models can be used in investigating the occurrence and progression of multiple vascular leak-driven diseases.The pre-clinical and clinical studies of TCM for promoting blood circulation and removing blood stasis provide fundamental support for the application of the above-mentioned platforms,with the purpose of uncovering the scientific basis of TCM for promoting blood circulation and removing blood stasis.

Key words：traditional Chinese medicine;blood circulation;blood stasis;tumor vessels;cerebral cavernous malfor⁃mation;diabetic retinopathy

Corresponding author：ZHAO Yang,E-mail：y.zhao@njucm.edu.cn;LU Yin,E-mail：luyingreen@njucm.edu.cn

血小板增多对小鼠恶性黑色素瘤转移模型的影响及机制

陈芋屹，周 杰，张甘霖，杨国旺

（首都医科大学附属北京中医医院肿瘤科，北京 100010）

摘要：目的建立小鼠高血小板恶性黑色素瘤转移模型，为研究肿瘤细胞诱导血小板聚集（TCIPA）的体内现象及机制提供稳定的操作平台；探索血小板增多对小鼠恶性黑色素瘤转移模型中肺转移灶、原位瘤、其他脏器转移灶及肿瘤免疫微环境的影响，为临床转移性恶性黑色素瘤的发病机制提供基础研究证据。方法提取鼠源性洗涤血小板。采用7～8周龄雄性C57BL6小鼠，分为高血小板荷瘤组、荷瘤组和对照组，每组8只。高血小板荷瘤组经尾静脉注射洗涤血小板悬液和鼠源性恶性黑色素瘤B16F10细胞悬液，并经原位注射B16F10细胞悬液建立小鼠高血小板恶性黑色素瘤转移模型；荷瘤组在高血小板荷瘤组基础上将洗涤血小板悬液更换为同体积Tyrode′s液；对照组为注射同体积PBS和Tyrode′s液的健康小鼠。比较组间肺重、肺转移灶数量、其他脏器转移灶数量、原位瘤重等指标，探索血小板增多对模型特征的影响。通过流式细胞术检测血小板增多对荷瘤模型肺、脾、肿瘤组织内T细胞、巨噬细胞、NK细胞数量及亚型的影响。通过免疫组化检测血小板增多对肺内M2型巨噬细胞浸润及PDL1表达的影响。结果接种后第14天取材，与对照组比较，荷瘤组、高血小板荷瘤组小鼠均出现肺转移，且高血小板荷瘤组较荷瘤组小鼠肺重增大（162±17 vs 237±66 vs 358±64），肺转移灶数量增多（309±116 vs 776±373），差异具有统计学意义（P＜0.05）。此外，与荷瘤组比较，高血小板荷瘤组小鼠肾、肝和结肠转移灶数量均增多，但原位瘤重降低。流式细胞术结果显示，与对照组、荷瘤组比较，高血小板荷瘤组小鼠肺中F4/80+CD206+巨噬细胞比例增高（P＜0.05），但在脾和原位瘤内比例降低。CD4+T细胞、CD8+T细胞、FoxP3+Treg细胞、NK细胞、NKT细胞组间差异无统计学意义。免疫组化结果显示，高血小板荷瘤组小鼠肺内F4/80，CD206和PDL1表达均高于荷瘤组（P＜0.05）。结论血小板增多促进小鼠恶性黑色素瘤发生肺转移，其机制与血小板参与调节宿主巨噬细胞分布，上调肺转移灶内M2型巨噬细胞及PDL1表达相关。

关键词：恶性黑色素瘤；血小板增多；肿瘤转移

基金项目：国家自然科学基金（81774039；82174453）

通讯作者：张甘霖，E-mail：kalinezhang@163.com；杨国旺，E-mail：guowang_yang@163.com

基于TRPV1探讨辣椒素促进胃癌转移机制

邓 蕊1,2，余苏云1,2，成 鹏1,2，韦忠红1,2，赵 杨1,2，李晓曼1,2，吴媛媛1,2，王爱云1,2，陈文星1,2，陆 茵1,2,3

（1.南京中医药大学药学院，江苏 南京 210023；2.江苏省中药药效与安全性评价重点实验室，江苏 南京 210023；3.江苏省中医药防治肿瘤协同创新中心，江苏 南京 210023）

摘要：目的探究辣椒素受体（TRPV1）在辣椒素介导的胃癌转移过程中对胃癌细胞转移及运动能力的作用及其机制。方法①利用荧光素酶标记的胃癌细胞构建胃原位及尾静脉转移瘤模型，通过小动物成像仪在整体动物水平观察辣椒素对胃癌转移的影响；②通过划痕、Transwell及3D侵袭实验，在细胞层面检测辣椒素对胃癌细胞迁移及侵袭能力的影响；③结合KM plotter生存曲线分析、免疫荧光、q-PCR和Western印迹法验证TRPV1在辣椒素介导的胃癌转移过程中的作用；④利用CRISPR-Cas9技术敲除胃癌细胞TRPV1基因，通过体内外实验观察TRPV1的缺失对辣椒素促胃癌转移作用的影响；⑤长时间动态活细胞观察成像仪结合细胞示踪软件观察胃癌细胞运动能力；⑥微流控芯片检测技术观察胃癌细胞变性能力。结果①与模型对照组相比，辣椒素100 mg·kg-1组小鼠胃癌转移率及肝转移结节数显著增加；②与溶剂对照组相比，辣椒素4，8和16 μmol·L-1处理后，胃癌细胞迁移、侵袭及运动能力显著增加，且呈剂量依赖性；③TRPV1在胃癌中异常高表达，且与胃癌患者生存期呈显著负相关。辣椒素能够促进TRPV1 mRNA及蛋白水平表达，同时促进TRPV1下游相关蛋白表达；④胃癌细胞中TRPV1基因的缺失导致辣椒素介导的胃癌转移增加过程被显著抑制；⑤辣椒素促进胃癌细胞运动能力及变形能力增加，TRPV1的表达缺失使辣椒素的促胃癌细胞运动能力及变形能力被显著削弱。结论辣椒素通过促进胃癌细胞上TRPV1蛋白表达及功能发挥，促进胃癌细胞变形及运动能力增加，进而加速胃癌转移过程。

关键词：辣椒素受体；胃癌；运动能力；变形能力

基金项目：国家自然科学基金（81973734；82004124）；南京中医药大学中药一级学科开放项目（2020YLXK20）

通讯作者：陆 茵，E-mail：luyingreen@126.com

苦味中药激活胃肠道苦味受体在不同疾病中的应用

顾任钧1，2，夏雅雯1，3，4，任 浪1，2，陶睿智1，3，4，陆可沁1，3，4，陆 茵1，3，4，孙志广1，2

（1.南京中医药大学，江苏 南京 210023；2.南京中医药大学附属江苏省第二中医院，江苏 南京 210019；3.南京中医药大学药学院江苏省中药药效与安全性评价重点实验室，江苏 南京 210023；4.南京中医药大学江苏省中医药防治肿瘤协同创新中心，江苏 南京 210023）

摘要：苦味受体（TAS2R）是一类7次跨膜的G蛋白偶联受体，它不但分布在味觉系统，还广泛分布在消化道系统。大多毒物或有害食物味苦，苦味感受器越敏感，越能调动人体的自我保护机制，物种规避毒物的能力和生存概率就越大。当机体遭遇疾病侵袭时，胃肠道TAS2R会激活并启动防御机制，进而影响为胃肠道及内分泌系统的功能。在中医药的应用中早就存在中药药性理论、中药五味理论以及系统的苦味中药功效，苦味作为五味之一，被广泛应用到各类疾病的治疗中。本文综述了苦味中药激活胃肠道苦味受体对不同疾病的防治作用，为苦味中药的防治机理提供新视角。

关键词：苦味受体；中医；中药；胃肠道

通讯作者：孙志广，E-mail：sunzhiguang@njucm.edu.cn

三七皂苷-3体外抗登革2型病毒作用

何雪梅，刘俊珊，曹惠慧，卢子滨，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的研究三七皂苷-3（简称三七-3）体外抗登革2型病毒（DENV-2）的作用。方法采用DENV-2新几内亚株感染BHK-21细胞，通过CCK-8法评价三七-3对DENV-2感染细胞的保护作用；噬斑法检测DENV-2感染细胞后子代病毒的合成；实时荧光定量PCR法和Western印迹法检测病毒结构蛋白E和非结构蛋白NS1的mRNA和蛋白表达水平；免疫荧光法检测病毒复制中间体dsRNA以及E和NS1蛋白的合成，并通过时间点处理实验判断抗DENV-2的作用阶段。结果三七-3能保护DENV-2诱导的细胞死亡，其半数有效浓度为6.314 μmol·L-1，抑制DENV-2诱导的子代病毒颗粒的释放，降低E和NS1的mRNA和蛋白表达水平，并抑制病毒dsRNA的合成，其抑制作用主要发生在病毒复制的早期阶段。结论三七-3具有显著的体外抗DENV-2的作用，有望成为抗登革病毒的候选药物。

关键词：三七皂苷-3；登革病毒；抗病毒作用；病毒复制

通讯作者：余林中，E-mail：yulzh@smu.edu.cn

杜仲叶黄酮对成骨细胞MG-63增殖的抑制作用及其机制

何周蔓，遆慧慧

（广东药科大学临床药学院，广东 广州 510006）

摘要：目的提取分离杜仲叶黄酮类物质，阐明其对体外培养的人成骨MG-63细胞的影响及其可能的分子机制。方法采用超声-微波协同法提取杜仲叶黄酮类物质；UPLC/Q-TOF-MS分析黄酮类物质单体；采用杜仲叶黄酮0～60 g·L-1及成骨诱导剂处理人成骨MG-63细胞48 h，MTT法分析其对细胞存活率的影响；采用杜仲叶黄酮7.5～60 mg·L-1协同成骨诱导剂处理人成骨MG-63细胞48 h，Western印迹法分析各组细胞中TLR-4，MyD88和NF-κB蛋白的表达水平。结果杜仲中主要含有黄酮类物质单体芦丁和槲皮素。杜仲叶黄酮可显著抑制MG-63细胞的增殖，且具有浓度依赖性，IC50为46.32 mg·L-1；升高MG-63细胞中TLR-4，MyD88和NF-κB蛋白的表达。结论杜仲叶黄酮类物质对成骨细胞MG-63增殖的抑制作用可能与激活TLR-4，MyD88和NF-κB等免疫分子信号通路相关。

关键词：杜仲；黄酮；TLR-4；成骨MG-63细胞

基金项目：国家自然科学基金（81873071）；广州市珠海科技新星（2019060610059）

通讯作者：遆慧慧，E-mail：tihuihui@126.com

降香及其替代药材改善急性心肌缺血所致的心肌损伤及冠脉微循环障碍

李 安1，汪文龙1，陈兰英1，欧阳灿玥1，赖潇筱1，崔亚茹1，张 妮2，罗颖颖1，刘荣华2

（1.江西中医药大学中药固体制剂制造技术国家工程研究中心，江西 南昌 330006；2.江西中医药大学药学院，江西 南昌 330004）

摘要：目的探讨降香及替代药材交趾黄檀对急性心肌缺血所致的心肌损伤及冠脉微循环障碍的保护作用。方法采用左前降支冠脉结扎法对大鼠进行造模，造模后的大鼠随机分到模型组（MI）、尼可地尔组（3 mg·kg-1）、血塞通组（30 mg·kg-1）、降香水提物组（1.5，3和6 g·kg-1）、交趾水提物组（1.5，3和6 g·kg-1）及假手术组，造模24 h后给药，每天给药1次，连续给药10 d，末次给药1 h后进行观察及取材，假手术和MI组给予相应剂量的蒸馏水。RGB色度分析检测耳廓，足底部“血瘀证”；插管检测大鼠血流动力学指标动脉收缩压（SBP）、动脉舒张压（DBP）、左室收缩压（LVSP）、左室舒张末期压（LVEDP）和等容收缩期左室内压力的上升/下降的最大速率（±dp/dtmax）；试剂盒检测血清中谷草转氨酶（GOT）、肌酸激酶（CK）、肌酸激酶同工酶（CK-MB）和一氧化氮（NO）含量；HE染色进行组织病理学分析；ELISA检测血浆内皮细胞蛋白C受体（sEPCR）和血友病因子（vWF）含量；1，1′-双十二烷基-3，3，3′，3′-四甲基吲哚菁高氯酸盐〔DiI（C12）〕灌注检测梗死边界及梗死远端区微血管面积、密度、空隙度，并根据泊肃叶公式计算出微血管血流阻力；免疫组化法检测心肌组织中CD31的表达；RT-PCR检测一氧化氮合酶（eNOS）、血管生成素2（Angpt-2）、碱性成纤维细胞生长因子（b-FGF）、血小板衍生生长因子B（PDGF-B）、内皮生长因子（VEGF）和内皮生长因子受体2（VEGFR2）mRNA的表达。结果降香、交趾水提物可改善冠脉结扎所引起耳廓和足底RGB色度；提高大鼠SBP，DBP，LVSP和±dp/dtmax，改善心功能；可减轻梗死区心肌细胞萎缩，减小细胞间隙，减少炎症细胞浸润，减轻肌丝溶解；降香、交趾水提物能降低急性心肌梗死大鼠血清中GOT，CK和CK-MB及血浆中sEPCR和vWF的含量，可减轻心肌和内皮损伤，提高血浆NO浓度，改善内皮功能；降香、交趾水提物可显著增加梗死边界及远端区微血血管面积和密度；降低梗死边界和远端区微血管空隙度及血流阻力，可显著增加梗死边界CD31的表达；可增加促血管生成因子b-FGF，PDGF-B，Angpt-2，VEGF和VEGFR2 mRNA的表达。结论降香、交趾水提物均可改善急性心肌梗死引起的心功能及微血管障碍，可能是通过促进冠脉微血管再生发挥作用，说明降香和交趾黄檀在心肌缺血方面具有一定的替代性。

关键词：左前降支冠脉结扎；心肌损伤；冠脉微循环障碍；微血管再生；降香；交趾黄檀

基金项目：国家重点研发计划（2018YFC1706102）

通讯作者：陈兰英，E-mail：clyxy2513@163.com；刘荣华，E-mail：rhliu@163.com

中药调控谷氨酰胺代谢重编程在治疗肺癌中的潜力

李静阳1，向 丽2，孟宪丽2

（成都中医药大学1.药学院，2.中医药创新研究院，四川 成都 611137）

摘要：肺癌已成为全球癌症致死率最高的疾病，严重威胁人类的健康，因此对抗肺癌药物的研究与开发迫在眉睫。近年来研究表明，肺肿瘤细胞的代谢方式明显不同于正常细胞，其中，肺癌细胞表现出对谷氨酰胺较高的成瘾性。因此，谷氨酰胺的代谢重编程很可能对肺肿瘤微环境具有重要的调控作用。中药具有多途径、多靶点、多通路调控的特点，其能否通过调节谷氨酰胺的代谢重编程从而调控肺肿瘤细胞代谢微环境，进而发挥抗肺癌的作用值得探索。本文通过对临床常用抗肺癌中药（百部、白芍、斑蝥、柴胡、西洋参、桂枝、党参等）作用机制的文献分析并结合网络药理学研究发现，以上6味中药与肺癌治疗靶点密切相关；特别是，这些中药所含成分可与谷氨酰胺代谢通路的谷氨酰胺合成酶（GLUL）和谷氨酰胺酶（GLS）密切关联，提示中药在调控谷氨酰胺代谢重编程治疗肺癌中的潜力。本文综述了肺癌中谷氨酰胺代谢通路的分子机制，探讨了中药调控谷氨酰胺代谢重编程在治疗肺癌中的潜力，以期为靶向谷氨酰胺代谢从而抗肺癌的治疗策略提供一定的理论基础。

关键词：中药；肺癌；谷氨酰胺代谢；重编程

通讯作者：向 丽，E-mail：xianglydr@cdutcm.edu.cn；孟宪丽，E-mail：xlm999@cdutcm.edu.cn

复方中药塞络通对多发性脑梗死小鼠神经发生与神经修复作用及对Notch1/TLR4信号通路的调节机制

李珺媛＊，姚明江＊，范晓迪，王光蕊，徐 立，杨 斌，宋文婷，刘建勋

（中国中医科学院西苑医院基础医学研究所，中药药理北京市重点实验室，北京 100091）

摘要：目的初步探讨复方中药塞络通（SLT）对多发性脑梗死损伤小鼠的神经修复作用及其可能的作用机制。方法C57/B16小鼠随机分为假手术组、脑梗死模型组、阳性药丁苯酞（78 mg·kg-1）组、SLT（11，22和44）mg·kg-1组，除假手术组外，其他每组小鼠建立微球致多发性脑梗死模型。按相应剂量灌胃给药7 d，同时ip给予Brdu（50 mg·kg-1）7 d，每天对小鼠进行称重和神经功能缺陷评分，7 d后检测各组小鼠抓力，激光散斑多普勒法测定梗死半脑脑血流量，HE染色观察脑组织病理变化，免疫荧光单标及双标染色检测脑组织中NeuroD1，Brdu/DCX和Brdu/PAX6表达情况，ELISA法检测患侧大脑皮质组织BDNF和bFGF含量以及血清CXCL2，IL-17A和MCP-1含量。采用Western印迹法和qRT-PCR法测定患侧海马组织Notch1、中白细胞介素-1β（IL-1β）、肿瘤坏死因子α（TNF-α）、细胞Toll样受体4（TLR4）、髓样分化因子-88（MYD88）、核因子-κB p65（NF-κB p65）的蛋白和mRNA表达水平。结果与正常对照组比较，模型组小鼠神经行为评分升高，体重减轻，抓力降低，脑血流量降低，HE染色观察脑组织病变区域可见明显的神经细胞无序排列、坏死、肿胀及凋亡。海马齿状回区NeuroD1阳性细胞数增加，Brdu+/DCX+和Brdu+/PAX6+细胞数量增加。大脑皮质BDNF和bFgF含量减少，血清CXCL2，IL17A和MCP-1含量升高，海马组织Notch1，IL-1β，TNF-α，TLR4，MYD88和NF-κB p65的蛋白和mRNA表达水平升高（P＜0.05）。与模型组相比，各给药组小鼠神经功能缺陷评分降低，体重升高，抓力升高，脑血流量有所恢复，脑组织病变区域的细胞无序状态、坏死、肿胀及凋亡现象减轻。海马齿状回区NeuroD1阳性细胞数进一步增加，Brdu+/DCX+、Brdu+/PAX6+细胞数量进一步增加。大脑皮质BDNF和bFgF含量增加，血清CXCL2，IL-17A和MCP-1含量降低，海马组织Notch1，IL-1β，TNF-α，TLR4，MYD88和NF-κB p65的蛋白和mRNA表达水平降低（P＜0.05）。结论塞络通能明显改善多发性脑梗死小鼠神经功能与病理形态，其机制可能是通过抑制Notch1/TLR4信号通路，从而发挥促神经发生与神经修复作用。

关键词：塞络通；缺血性脑损伤；神经修复；炎症反应；Notch1/TLR4信号通路

基金项目：国家自然科学基金（81873040）；广西中医基础研究重点实验室开放课题（19-050-45-10）

通讯作者：姚明江，E-mail：ymj_ren@163.com；刘建勋，E-mail：liujx0324@sina.com

＊共同第一作者.

Chamaejasmine B通过调节细胞乳腺癌自噬降低血管内皮细胞对VEGFA促血管生成信号的敏感性

李 琦，阚晓溪，尹 婕，孙立东，王娅杰，李玉洁，杨 庆，肖红斌，陈 颖，翁小刚，蔡维艳，朱晓新

（中国中医科学院中药研究所，北京 100700）

摘要：目的探究瑞香狼毒提取物瑞香B（chamaejasmine B，ICJ）抗乳腺癌的药效及其作用机制。根据前期研究提示，重点聚焦其对乳腺癌血管生成的影响。方法通过原位乳腺癌模型探究ICJ抗乳腺癌的药效；通过基质凝胶塞实验、Tran⁃swell、内皮细胞毛细血管成管实验和主动脉环实验，进一步观察ICJ对于乳腺癌血管生成的影响，通过透射电镜观察ICJ对于细胞自噬的影响；免疫共沉淀、Western印迹法和RT-PCR分别检测相关蛋白和mRNA表达。结果通过原位乳腺癌模型，我们首次证明了ICJ对乳腺癌具有显著抗血管生成作用。基质凝胶塞实验进一步验证了ICJ的体内疗效。基于共培养模型，通过Transwell、成管和主动脉环实验，证明ICJ有效抑制肿瘤-HUVEC的新生血管。在机制层面，ICJ对HUVEC细胞凋亡影响较小，LC3Ⅰ/Ⅱ比值升高，LC3点状分布，Beclin-1上调，明显诱导自噬。此外，免疫共沉淀实验证明，ICJ通过与LC3结合，进而抑制VEGFR2水平，其抗血管生成效果与自噬的发生密切相关。结论ICJ可通过VEGFR2减弱促血管生成信号，是一种新型的血管生成抑制剂，有潜力成为乳腺癌抗血管生成化疗的补充用药。

关键词：chamaejasmine B；抗血管生成；化疗疗法；乳腺癌；自噬

基金项目：国家科技重大专项（2017ZX09101002-002-002）；中国中医科学院优秀青年科技人才专项培养计划（ZZ13-YQ-044）；中国中医科学院“一带一路”合作项目〔（20141202）GH201914〕；中药研究所内部项目（Z2017019-03）；国家自然科学基金（81303272）

通讯作者：朱晓新，E-mail：zhuxx@icmm.ac.cn

四妙方通过调节肝脏代谢和肠道菌群改善非酒精性脂肪性肝病

李 艳，韩瑞婷，盛丽莉，李后开

（上海中医药大学交叉科学研究院，上海 201203）

摘要：目的非酒精性脂肪性肝病（NAFLD）是目前全世界最为常见的肝病，而高尿酸是NAFLD的独立危险因素。痛风与NAFLD具有相同的证型—湿热蕴结和相同的病理基础—高尿酸。四妙方（SMF）是临床常用的经典中药方剂，通过降低血清尿酸水平治疗痛风和痹症。因此，本研究基于“异病同治”的中医治则，通过高脂高糖饮食诱导的小鼠模型，从尿酸、肝代谢和肠道菌群的角度研究四妙方对于NAFLD的改善作用。方法首先，采用高脂高糖饮食连续干预16周诱导C57BL/6J小鼠NAFLD模型，同时ig给予四妙方16周，测定不同组别小鼠肝和血清中的脂质水平及尿酸水平，用组织病理学染色评价肝脂质积聚的程度。再通过肝转录组学富集差异表达基因，预测调控途径，采用实时荧光定量PCR技术（qRT-PCR）检测肝脂质代谢与炎症相关指标。最后，采用16S微生物多样性测序测定肠道微生物的组成。结果SMF可以显著改善高脂高糖饮食小鼠的肝脂肪变性、减缓体重增加、降低血清尿酸浓度，改善胰岛素的敏感性和葡萄糖耐量。其次，肝转录组学结果显示，SMF抑制了脂肪酸的生物合成并刺激了胰岛素的分泌。qRT-PCR结果显示，SMF下调了肝脂类代谢相关基因（Acly，Fas，Acc和Scd-1）和促炎细胞因子（IL-1β和Nlrp-3）的表达，与肝转录组学结果一致。最后，SMF显著改变了小鼠的肠道菌群组成，特别是增加了Akkermansia muciniphila的丰度。结论SMF可以通过调节肝脂质代谢途径，及调节肠道菌群组成，特别是增加Akker⁃mansia muciniphila的丰度，减轻高脂高糖饮食引起的NAFLD。

关键词：四妙方；非酒精性脂肪性肝病；肝代谢；肠道菌群

基金项目：上海市卫生健康委员会中医药研究计划（2020JP016）；国家重点研发计划（2017YFC1700200）

通讯作者：李后开，E-mail：houkai1976@126.com

BH3拟似药物的发现及与组蛋白去乙酰化酶抑制剂的协同抗肿瘤作用

李玉倩1，李学军2

（1.石河子大学药学院，新疆 石河子 832000；2.北京大学医学部基础医学院，北京 100191）

摘要：目的BH3-only蛋白是Bcl-2家族的促凋亡成员，可与抗凋亡成员结合启动肿瘤细胞凋亡和自噬。BH3拟似药通过模拟BH3-only蛋白促进肿瘤细胞凋亡和自噬，是肿瘤治疗中最具前景的药物之一。因此我们的研究目的是寻找新的BH3拟似药并阐明其抗肿瘤作用机制，为发展新的BH3拟似药奠定基础。方法通过基于药效团模型的虚拟筛选技术和分子对接筛选出潜在的BH3拟似化合物。用MTS实验检测筛选出来的化合物对多种肿瘤细胞的抑制率。采用Annexin V/PI双染流式细胞术检测化合物对细胞凋亡的影响，免疫共沉淀（Co-IP）检测化合物对Bcl-2/Beclin-1复合物的影响。共聚焦显微镜检测化合物对肿瘤细胞自噬的影响，采用Western印迹法检测凋亡和自噬相关蛋白的表达。通过hoechst 33342染色、透射电镜、MDC染色检测AD23联合伏立诺他对肿瘤细胞蛋白与自噬的影响，动物实验检测AD23联合伏立诺他的抗肿瘤效果。结果筛选出BH3拟似化合物AD23。分子对接和表面等离子共振实验结果表明，AD23可与Bcl-2，MCL-1和Bcl-XL蛋白结合，通过MTS实验发现AD23抑制小细胞肺癌细胞系作用最强。Annexin V/PI双染实验结果显示，AD23可以浓度依赖性的促进H82细胞凋亡。Western印迹实验结果显示，AD23可以剂量依赖性诱导胱天蛋白酶3活化，诱导C-PARP蛋白增多从而激活凋亡通路。mCherry-GFP-LC3质粒转染结果显示，AD23可以浓度依赖性的促进自噬流过程的发生。Co-IP实验结果表明，AD23可时间依赖性促进Bcl-2与Beclin-1解离，AD23可以浓度依赖性的上调LC3Ⅱ/Ⅰ蛋白比例增加，下调P62蛋白的表达，从而激活自噬通路。细胞及动物实验结果表明AD23联合伏立诺他给药组促进H82细胞凋亡与自噬较单独给药组明显增加。结论BH3天然拟似化合物AD23可促进小细胞肺癌细胞凋亡与自噬，AD23与伏立诺他具有协同抗肿瘤作用。

关键词：肿瘤；BH3天然拟似化合物；凋亡；自噬；组蛋白去乙酰化酶抑制剂

小半夏汤通过调节Gasdermin E改善大鼠化疗性异食癖

廖秀秀，聂 克

（广东药科大学中药学院，广东 广州 510006）

摘要：目的建立化疗性异食癖大鼠模型，观察小半夏汤对化疗大鼠Gasdermin E（GSDME）表达水平的影响，从GSDME介导的细胞焦亡角度探讨小半夏汤防治化疗性恶心呕吐的作用机制。方法将Wistar大鼠随机分为正常对照组、顺铂模型组、昂丹司琼（2.6 mg·kg-1）组和小半夏汤治疗（3.2 g·kg-1）组；首次ig给药1 h后，除正常对照组外，其余各组ip给予顺铂（6 mg·kg-1）建立大鼠化疗性异食癖模型，以大鼠高岭土摄食量作为其恶心呕吐程度指标，每24 h记录大鼠的高岭土摄食量。连续给药3 d，造模72 h后取血和胃窦。采用ELISA方法测定血清中IL-1β，IL-18，HMGB1含量；HE染色观察组织病理情况；实时荧光定量PCR测定各组大鼠胃窦GSDME的mRNA表达水平；Western印迹法测定大鼠胃窦胱天蛋白酶3、活化胱天蛋白酶3，GSDME和GSDME-N蛋白的表达。结果①与正常对照组比较，造模后24和72 h内模型组大鼠的高岭土累积摄食量均显著增加（P＜0.01），成功建立大鼠异食癖模型；与模型组比较，造模后24和72 h内小半夏汤治疗组和昂丹司琼治疗组高岭土摄食累积量均显著降低（P＜0.05，P＜0.01）。②HE染色结果显示，与正常对照组比较，顺铂模型组胃窦组织上皮表面严重破坏，固有层腺体排列疏松、紊乱，固有层腺体嗜酸性增强，可见明显炎性浸润。与顺铂模型组相比，昂丹司琼治疗组和小半夏汤治疗组可显著改善胃窦组织损伤，且小半夏汤治疗组损伤程度小于阳性药组，但不及正常对照组。③实时荧光定量PCR结果显示，与正常组比较，模型组大鼠胃窦GSDME mRNA表达显著升高（P＜0.01）；与模型组比较，小半夏汤治疗组胃窦GSDME mRNA表达显著降低（P＜0.01）。④Western印迹法结果显示，与正常组比较，模型组大鼠胃窦的活化胱天蛋白酶3、GSDME-N显著升高（P＜0.01）；与模型组比较，小半夏汤组胃窦的活化胱天蛋白酶3和GSDME-N显著降低（P＜0.01）。⑤ELISA结果显示，与正常组比较，模型组大鼠血清中IL-1β，IL-18和HMGB1含量显著升高（P＜0.01）；与模型组比较，小半夏汤组血清中IL-1β，IL-18和HMGB1含量均显著降低（P＜0.01）。结论小半夏汤可以通过抑制胃窦GSDME的表达和切割，减少化疗导致的胃窦组织细胞焦亡，从而改善化疗性恶心呕吐。

关键词：小半夏汤；Gasdermin E；细胞焦亡

基金项目：国家自然科学基金（82174143）

通讯作者：聂 克，E-mail：nicknk@hotmail.com

桑枝总生物碱片的守正创新之路

刘率男，刘 泉，刘玉玲，谢明智，申竹芳

（中国医学科学院北京协和医学院药物研究所，天然药物活性物质与功能国家重点实验室，药物传输技术及新型制剂北京市重点实验室，中国医学科学院糖尿病研究中心，北京 100050）

摘要：我国原创降血糖天然药物“桑枝总生物碱片”于2020年3月获批上市，是十几年来我国首个获批的中药创新药，也是糖尿病治疗领域唯一的有效组分天然药物。在其研发过程中，根据中药传统理论，并结合现代药理学研究技术方法，确定了桑枝中的降血糖作用有效组分。在临床前药理学过程中，采用了多种现代药理学研究方法，挖掘出桑枝总生物碱具有作用靶点清晰、物质基础明确和降血糖作用显著等特点；在临床研究中发现其广泛的糖尿病治疗综合获益远超单纯的糖苷酶抑制剂，可能具有天然药物独特的多重药理作用优势。因此，后续采用了多种现代药理学手段，广泛地挖掘桑枝总生物碱的多重药理作用特点与潜在机制。本文将就围绕其研发历程及后续机制探索工作，浅谈利用现代药理学研究理念，从天然药物宝库中获取有效活性成分和新药研究中需要突破的技术瓶颈和理念创新，以期为我国创新中药的创制工作提供研究实例经验参考。

关键词：桑枝总生物碱；2型糖尿病；餐后血糖

基金项目：北京市自然科学基金（7202137）；国家自然科学基金（81973379）；北京市高校卓越青年科学家计划（BJJW⁃ZYJH0120190023028）

通讯作者：申竹芳，E-mail：shenzhf@imm.ac.cn

海洋来源天然产物金黄霉素A抗神经炎症作用及机制

刘 漫，张姗姗，刘冬妮，杨滢霖，方莲花，王月华，杜冠华

（中国医学科学院北京协和医学院药物研究所，药物靶点研究与新药筛选北京市重点实验室，北京 100050）

摘要：目的研究海洋来源天然产物金黄霉素A（Chr-A）抗神经炎症作用及机制。方法ip给予LPS 5 mg·kg-1构建小鼠神经炎症模型，LPS 200 μg·L-1刺激BV2小胶质细胞构建炎症反应细胞模型。RNA-seq转录组分析小鼠脑皮质中的差异基因；通过Metascape在线平台（https：//metascape.org/）和String在线平台（https：//string-db.org/）对Chr-A调控的差异基因进行富集分析和PPI分析。ELISA法检测细胞炎症因子的含量；Western印迹法和RT-PCR检测炎症相关mRNA和蛋白的表达。结果RNA-seq结果进行差异基因（DEGs）分析（FC＞1.5，P＜0.05），发现LPS模型组与正常对照组相比较有639个DEGs，Chr-A给药组和LPS模型组相比较有113个DEGs，综合分析发现与Chr-A调控LPS诱导神经炎症相关的70个DEGs；GO富集分析表明Chr-A抗神经炎症可能与response to cytokine，cellular response to cytokine stimulus和regula⁃tion of immune system process等生物过程密切相关；KEGG富集分析表明Chr-A发挥抗神经炎症作用可能与涉及到TNF signaling pathway，cytokine-cytokine receptor interaction，NF-κB signaling pathway，IL-17 signaling pathway和 NOD-like receptor signaling pathway等信号通路。应用RT-PCR进行深入验证，发现小鼠脑皮质中TNF-α，IL-6，IL-1β，CCL2和NLRP3的mRNA表达水平的结果与RNA-seq一致。此外，Chr-A显著降低小鼠脑皮质中炎症因子IL-6，IL-1β，IL-17，TNF-α，GM-CSF，CXCL12和MCP-1的水平，抑制COX2、NLRP3和活化胱天蛋白酶1蛋白的表达。在LPS刺激BV2细胞炎症反应模型上，Chr-A显著降低细胞上清中NO，IL-6，IL-1β，TNF-α和MCP-1的含量，下调COX2、NLRP3和活化胱天蛋白酶1等蛋白的表达。结论Chr-A具有抗神经炎症作用，其机制可能与下调NLRP3/活化胱天蛋白酶1信号通路相关。

关键词：金黄霉素A；脂多糖；神经炎症；小胶质细胞

基金项目：国家重点研发计划（2018YFC0311005）

通讯作者：王月华，E-mail：walling@imm.ac.cn；杜冠华，E-mail：dugh@imm.ac.cn

生姜活性分子8GL在急性髓系白血病分化治疗中的应用

卢伟强，易春阳，赵玉淼，刘明耀

（华东师范大学生命科学学院，上海 200241）

摘要：目的急性髓系白血病（AML）是一种常见的恶性血液肿瘤，具有高侵袭性和高异质性的特征。分化诱导治疗在部分AML患者中取得了巨大成功，然而大多数AML患者仍无法从该疗法中获益，其5年生存率仍＜30%。新分化机制的探索和新靶向药物的开发依然是AML治疗的迫切需求。G蛋白偶联受体（GPCR）在多种生理和病理过程中发挥关键作用，是最重要的药物靶标家族之一。本研究聚焦于孤儿GPCR在AML细胞分化调控中的作用，以期为AML的治疗研究提供新的作用机制，新靶点和新药物。方法利用AML癌症基因组学数据挖掘潜在的孤儿GPCR靶标；利用体内外模型考察孤儿GPCR在白血病细胞分化调控中的作用；建立孤儿GPCR药物筛选模型，筛选中药单体化合物库，发现新的GPCR配体，考察其体内外促分化的作用及其分子机制。结果基于AML癌症基因组学分析发现，孤儿GPR-X特异性表达于髓系细胞，其高表达与AML的良好预后相关。通过生物实验，发现激活GPR-X在体内外均能显著促进AML细胞分化，抑制肿瘤细胞生长，说明GPR-X是一个新的AML细胞分化调节蛋白。通过Tango、NanoBit等多种孤儿GPCR药物筛选体系，鉴定出一种新型高效GPR-X激动剂——8GL。8GL来源于传统药用植物生姜，能够通过激活GPR-X促进AML细胞的分化。同时，8GL在体内仍具有诱导治疗效果且毒副作用较低，具有良好的安全性。机制研究表明，8GL在AML细胞中通过激活GPR-X-Gs-PKA通路阻断mTOR信号通路，进而发挥分化诱导作用。联用8GL与mTOR抑制剂依维莫司能够发挥协同抗癌作用。结论本研究发现了一个新的AML分化诱导受体GPR-X，并鉴定出一个全新的高效GPR-X激动剂8GL。8GL通过调控GPR-X下游的Gs-PKA-mTOR信号轴调控AML细胞分化，并与mTOR抑制剂发挥协同抗AML作用。

关键词：白血病；分化治疗；G蛋白偶联受体；激动剂

通讯作者：卢伟强，E-mail：wqlu@bio.ecnu.edu.cn

丹酚酸B促进乳腺癌小鼠肿瘤血管正常化

钱 程1，仲崇金3，韦忠红1，赵 杨1，3，陈文星1，2，王爱云1，2，陆 茵1，2

（南京中医药大学1.药学院，江苏省中药药效与安全性评价重点实验室，2.江苏省中医药防治肿瘤协同创新中心，3.医学院·整合医学学院，江苏 南京 210023）

摘要：目的中医辨证中，丹参属活血化淤药，能够治疗瘀血症候，肿瘤作为癥积，属久瘀所致。基于丹参在心肌缺血、冠心病等疾病中发挥改善血管结构和促进血管灌注的作用，而肿瘤血管也存在结构异常和灌注不足的特征，本研究着眼于丹参主要活性成分丹酚酸B对于肿瘤微血管的调控，是否能促进肿瘤血管正常化。方法选用健康雌性小鼠，注射乳腺癌细胞悬液，构建小鼠原位乳腺癌模型，分为模型组、丹酚酸B低剂量给药组、高剂量给药组。每3 d测定肿瘤体积。实验周期结束，每组选择小鼠进行尾静脉注射TRITC-Dextran和Pimonidazole，考察肿瘤血管渗漏以及缺氧状况。利用扫描电子显微镜，观察不同组别内部血管内皮内衬。使用免疫荧光染色，测定CD31/α-SMA、CD31/NG2、CD31/VE-cadherin、CD31/claudin5等，分析肿瘤血管结构。使用小动物光声成像系统，考察肿瘤组织内部的血氧灌注情况，评判肿瘤血管功能。结果与模型组相比，丹酚酸B低剂量和高剂量组均没有显著抑制肿瘤体积增长。免疫荧光结果显示，相比于模型组，丹酚酸B低剂量组和高剂量组能够显著的增加肿瘤血管成熟细胞覆盖率（P＜0.05），并增强内皮细胞间的紧密连接（P＜0.05）。CD31/TRITC-Dextran结果显示，丹酚酸B能够显著降低肿瘤血管的渗漏，能够促进肿瘤血管结构的完整。Pimonidazole免疫荧光结果显示，丹酚酸B低剂量组和高剂量组的血管缺氧情况得到显著改善（P＜0.05）。活体成像系统结果显示一致结果，丹酚酸B能够促进肿瘤组织内部的血氧灌注。结论丹酚酸B作为活血化瘀中药丹参中主要活性成分，能够通过促进肿瘤血管壁成熟细胞覆盖、加强肿瘤内皮细胞间紧密连接，从而改善血管渗漏，促进血管携氧功能。揭示其促进肿瘤血管正常化的作用。

关键词：丹酚酸B；乳腺癌；肿瘤血管正常化

基金项目：国家自然科学基金（81973587；81973734；82003991）；全国中医药创新骨干人才培训项目

通讯作者：陆 茵，E-mail：luyingreen@126.com；王爱云，E-mail：way9815@163.com

甘草正丁醇提取物抑制登革2型病毒的作用

石玲珠，刘俊珊，曹惠慧，卢子滨，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的对甘草正丁醇提取物抑制登革病毒（DENV-2）的活性进行了评价，并对其作用机制进行初步探讨。方法体外实验采用BHK-21细胞测定甘草正丁醇提取物对细胞的半数有毒浓度（CC50）以及对DENV-2的半数抑制浓度（IC50）；通过CCK-8法在DENV-2感染后不同时间点以及不同的感染阶段加入药物检测细胞存活率；通过噬斑实验检测子代病毒释放水平；通过qRT-PCR、Western印迹法和免疫荧光实验检测病毒E和NS1的表达。体内实验采用7日龄ICR乳小鼠，颅内加ip给予DENV-2建立全身性感染乳小鼠模型，分别于造模后的第1，3和5天ig给药进行治疗，每天观察并记录乳小鼠临床评分及生存率；采用qRT-PCR、Western印迹法和免疫荧光实验检测各组织病毒载量；通过ELISA检测脑、肝、脾炎症因子的表达；通过HE染色观察各组织病变。结果体外研究表明，甘草正丁醇提取物的CC50为（202.61±2.31）mg·L-1，IC50为（6.24±1.16）mg·L-1；细胞感染DENV-2的同时给予甘草正丁醇提取物可显著改善细胞死亡，减少子代病毒释放量并抑制病毒E和NS1蛋白表达。体内研究表明，甘草正丁醇提取物对感染DENV-2的ICR乳小鼠具有治疗作用，具体表现为临床评分改善，生存时间延长；同时乳小鼠组织病毒载量明显降低，有效调节病毒感染导致的炎症细胞因子白细胞介素6（IL-6）、肿瘤坏死因子α和IL-1β的变化，进而减轻组织病变。结论甘草正丁醇提取物在体内体外均能显著抑制DENV-2感染，其通过靶向DENV-2上的包膜蛋白抑制病毒复制周期的吸附过程进而发挥抗病毒作用。提示甘草正丁醇提取物可能是一种有前景的治疗登革病毒的候选药物。

关键词：甘草正丁醇提取物；登革病毒；抗病毒作用；吸附

通讯作者：余林中，E-mail：yulzh@smu.edu.com

人参皂苷元AD-1抗肺纤维化作用及机制

苏光悦，柯昂昂，李 涛，陈 钰，邹予文，贾俊丰，符美慧，赵余庆

（沈阳药科大学功能食品与葡萄酒学院，辽宁 沈阳 110016）

摘要：目的探讨人参皂苷元AD-1抗体内外肺纤维化的作用及其机制，为后疫情时代肺损伤保护作用的中药研发提供理论依据。方法体外实验利用脂多糖（LPS）10 mg·L-1刺激小鼠成纤维细胞L929 24 h，再分别给予AD-1 5，10和20 μmol·L-1进行治疗，采用MTT法检测AD-1对LPS诱导的成纤维细胞增值毒性的影响；采用流式细胞术检测AD-1对LPS诱导的L929细胞凋亡的影响；采用划痕愈合实验检测AD-1对L929细胞迁移的影响；采用Western印迹法检测AD-1对LPS诱导的L929细胞中α-平滑肌肌动蛋白（α-SMA）、Bax、Bcl-2、基质金属蛋白酶抑制剂1（TIMP-1）和基质金属蛋白酶2（MMP-2）的表达水平。在体实验采用昆明小鼠单次博来霉素（BLM，3.5 mg·kg-1）气管灌注的方法建立小鼠特发性肺纤维化模型。术后随机分为对照组（对照组气管灌注等体积的生理盐水），BLM组，BLM+AD-1（5，10和20 mg·kg-1）组，BLM+醋酸泼尼松（PA，5 mg·kg-1）组，于术后24 h灌胃治疗，连续给药28 d后取材，计算小鼠肺系数，并采用ELISA法检测小鼠肺组织羟脯氨酸（HYP）含量；采用Western印迹法检测α-SMA、转化生长因子β1（TGF-β1）、TIMP-1等纤维化因子的表达，并检测Bax、Bcl-2、β-连环蛋白和人沉默调节蛋白3（Sirt-3）等凋亡蛋白的表达。结果体外实验结果显示，AD-1可显著抑制LPS诱导的L929细胞增殖与迁移（P＜0.01）；与LPS（10 mg·L-1）组比，AD-1 5，10和20 μmol·L-1均可显著降低Bax/Bcl-2比值改善LPS诱导的L929细胞损伤引起的细胞凋亡（P＜0.01）；10和20 μmol·L-1组可显著增加MMP-2/TIMP-1比值（P＜0.01），提示AD-2可能通过促进细胞外基质的降解从而改善LPS诱导的L929细胞损伤。在体实验结果显示，与BLM模型组相比，AD-1 10和20 mg·kg-1均能显著降低BLM诱导的小鼠肺系数增加（P＜0.01）；显著降低血清及肺泡灌洗液中HYP的表达水平（P＜0.01），并体现出剂量依赖性；显著降低BLM诱导的α-SMA，TGF-β1及TIMP-1的表达增加（P＜0.05）；显著降低Bax/Bcl-2，p-P53和β-连环蛋白表达（P＜0.01）；显著增加Sirt3表达水平（P＜0.01），提示AD-1对BLM所致小鼠肺纤维化具有抑制作用。结论AD-1能够改善LPS诱导的成纤维细胞损伤及BLM诱导的小鼠肺纤维化，机制可能与改善正常肺细胞凋亡及抑制TGF-β1/α-SMA/Sirt3信号通路有关。

关键词：人参皂苷元AD-1；肺纤维化；L929细胞；细胞凋亡

基金项目：国家自然科学基金（81703386）；沈阳药科大学骨干人才培育计划（ZQN2018003）

通讯作者：赵余庆，E-mail：zyq4885@126.com

颈动脉粥样硬化患者单核细胞亚群、血脂水平与血管内皮功能关联分析

谭展飞1，张业昊1，李 浩2，刘建勋1

（1.中国中医科学院西苑医院基础医学研究所，中药药理北京市重点实验室，北京 100091；2.中国中医科学院望京医院，北京 100102）

摘要：目的探讨不同程度颈动脉粥样硬化患者外周血单核细胞亚群、血脂水平与血管内皮功能的相关性，为该病的发病与治疗提供新思路。方法以2020年9月至2021年3月期间在中国中医科学院西苑医院门诊招募的66例颈动脉粥样硬化患者为研究对象，以同期52例排除颈动脉粥样硬化的健康受试者为对照组。采用流式细胞仪检测各组血清单核细胞亚群（CD14++CD16-“经典型-Mon1”，CD14++CD16+“中间型-Mon2”和CD14+CD16++“非经典型-Mon3”）水平；采用生化仪检测血脂〔总胆固醇（CHO）、甘油三酯（TG）、高密度脂蛋白胆固醇（HDL-C）、低密度脂蛋白胆固醇（LDL-C）、脂蛋白a、极低密度脂蛋白胆固醇（VLDL-C）〕水平；另采用彩色多普勒超声检查仪检测各组患者肱动脉内皮依赖性功能（FMD）和颈动脉内膜中层厚度（IMT）等血管内皮功能参数。采用Pearson相关性分析单核细胞亚群、血脂水平与血管内皮功能之间的相关性。结果颈动脉粥样硬化患者血清单核细胞亚群（CD14++CD16-、CD14++CD16+、CD14+CD16++）表达水平显著高于健康对照者（P＜0.05），且随着颈动脉粥样硬化程度加剧，血清单核细胞亚群表达水平逐渐上升；颈动脉粥样硬化患者血脂CHO、TG、LDL-C水平明显高于对照组（P＜0.05），且随着颈动脉粥样硬化程度的增加，血脂异常水平升高（P＜0.05）；颈动脉粥样硬化患者FMD值明显低于对照组（P＜0.05），IMT显著高于对照组（P＜0.05）。且随着颈动脉粥样硬化程度增加，患者血管功能各指标明显下降（P＜0.05）；Pearson相关性分析结果表明，血脂LDL-C、VLDL-C水平和单核细胞CD14++CD16+“中间型-Mon2”水平与血管内皮功能指标FMD水平呈显著负相关。结论颈动脉粥样硬化患者血管内皮功能下降，血脂水平、单核细胞亚群表达水平均与健康人存在差异表达，且表达水平与颈动脉粥样硬化严重程度以及血管内皮功能存在显著相关性。表明机体脂代谢和免疫反应机制参与颈动脉粥样硬化发生和发展，为颈动脉粥样硬化性免疫提供理论基础，为中医药防治动脉粥样硬化提供新思路。

关键词：颈动脉粥样硬化；单核细胞亚群；血脂水平；血管内皮功能

基金项目：国家自然科学基金（81873041）

通讯作者：李 浩，E-mail：xyhplihao1965@126.com；刘建勋，E-mail：liujx0324@sina.com

基于“心-肠”轴理论对动脉粥样硬化患者肠道菌群特征的研究

谭展飞1，张业昊1，李 浩2，刘建勋1

（1.中国中医科学院西苑医院基础医学研究所，中药药理北京市重点实验室，北京 100091；2.中国中医科学院望京医院，北京 100102）

摘要：目的本研究旨在基于“心-肠”轴理论研究动脉粥样硬化（AS）患者肠道菌群的表达特点，为中医药从肠道菌群抗AS提供参考依据和新思路。方法顺序选取2020年9月—2021年3月于西苑医院心血管科就诊的颈动脉粥样硬化患者34例作为动脉粥样硬化组（AS组），来院行健康体检的健康者30例作为健康对照组（NOR组）。采集2组人群的一般资料；行颈动脉彩超检查颈动脉内-中膜厚度和最大斑块面积；空腹抽血检测血脂、血糖、肝肾功等血液生化指标；采集粪便行16S rRNA扩增子测序，对比分析2组粪便菌群的丰度以及结构差异，并将丰度差异显著的属水平菌群与宿主颈动脉超声和血液生化指标进行相关性分析。结果对比2组受试者的年龄和性别，差异均无统计学意义，2组之间具有可比性。AS组的总胆固醇、甘油三酯、低密度脂蛋白、脂蛋白a水平显著高于非AS组（P＜0.05）。与NOR组相比，AS组肠道菌群的Simpson指数和Shanno指数提示2组间的α多样性未见显著差异，PCoA分析和NMDS分析均显示2组的肠道菌群β多样性有明显差异（P＜0.05）。与NOR组相比，AS组肠道菌群序列门水平拟杆菌门丰度明显降低（P＜0.05），厚壁菌门丰度明显升高（P＜0.05）。与NOR组相比，AS组肠道菌群序列属水平的布劳蒂亚菌属、小球菌属、梭菌属、梭杆菌属、厌氧菌属、梭形杆菌属、多雷亚菌属丰度明显升高（P＜0.05），普雷沃氏菌属、副拟杆菌属、萨特雷亚菌属丰度明显降低（P＜0.05）；布劳蒂亚菌属丰度与最大斑块面积、低密度脂蛋白水平具有显著正相关性（P＜0.05）；普雷沃氏菌属丰度与最大斑块面积、低密度脂蛋白水平具显著负相关性（P＜0.05）。结论AS患者的肠道菌群多样性与健康人群存在差异，肠道菌群中布劳蒂亚菌属和普雷沃氏菌属增加或可作为AS的特征生物学指标。未来开发针对特定肠内菌种与菌内蛋白为靶点的中药新药，采用多靶点、多功效、取材天然、安全性高的中药复方制剂或可为AS的防治带来新的希望。

关键词：肠道菌群；动脉粥样硬化；“心-肠”轴；临床特征；16S rRNA扩增子测序

基金项目：国家自然科学基金（81873041）

通讯作者：李 浩，E-mail：xyhplihao1965@126.com；刘建勋，E-mail：liujx0324@sina.com

大波斯菊苷通过诱导乳腺癌细胞自噬抑制乳腺癌的发展

汪 丹，潘 燕

（北京大学基础医学院药理学系，北京 100191）

摘要：目的乳腺癌是威胁全球女性健康排名第一的恶性肿瘤，全球每年新增约226万患者。尽管随着诊断和治疗手段的发展，乳腺癌患者的生存期得到了显著延长，但是仍有每年超过50万的乳腺癌患者死亡。因此，有效遏制乳腺癌以及探索针对乳腺癌的治疗靶点成为迫切需要。大波斯菊苷（cosmosiin），又名芹菜素-7-葡萄糖苷（apigenin-7-glucoside），是芹菜素（apigenin）的糖苷衍生物，属于黄酮苷类化合物，广泛分布于洋蓟、洋甘菊和橄榄等植物中。据报导，大波斯菊苷具有抗糖尿病和治疗阿尔茨海默病的药理活性。此外，有研究发现，大波斯菊苷具有抗肿瘤的能力，但是其机制尚不明确，并且其对于乳腺癌的药理作用尚无报导。方法利用MTS法检测大波斯菊苷对乳腺癌MCF-7细胞体外增殖能力的影响；利用Transwell试验检测MCF-7细胞的迁移能力变化；利用纤维粘连蛋白包被的96孔板检测细胞黏附能力；Western印迹法实验检测自噬以及代谢相关分子的蛋白表达水平的变化。结果大波斯菊苷7.5～30 mg·L-1能剂量依赖地抑制MCF-7细胞的增殖、迁移和黏附能力。此外，大波斯菊苷30 mg·L-1处理MCF-7细胞24 h后，细胞中P62蛋白表达水平显著下调，LC3BⅡ/LC3BⅠ蛋白表达水平的比值显著升高，且p-AMPK的蛋白表达水平显著上调。结论大波斯菊苷可能通过激活AMPK信号通路促进乳腺癌细胞自噬，从而抑制乳腺癌的发展。

关键词：大波斯菊苷；乳腺癌；自噬

通讯作者：潘 燕，E-mail：pannay26@bjmu.edu.cn

大黄蒽醌类成分影响肠道微生物的组成平衡损伤结肠黏膜屏障促进结肠癌发展

韦忠红1,2，赵 杨1,2，李晓曼1,2，吴媛媛1,2，陈文星1,2，王爱云1,2，陆 茵1,2,3

（1.南京中医药大学药学院，江苏 南京 210023；2.江苏省中药药效与安全性评价重点实验室，江苏 南京 210023；3.江苏省中医药防治肿瘤协同创新中心，江苏 南京 210023）

摘要：目的蒽醌类泻药的安全性一直存在争议。此前，一些碎片化的研究发现蒽醌类药物可破坏结肠上皮组织结构，长期服用蒽醌类泻药可增加结直肠癌的风险。本研究以蒽醌类泻药中的主要通便成分sennoside A为研究对象，以肠黏膜屏障的完整性为切入点，着眼于肠道微生物所扮演的角色，研究sennoside A对肠道炎症和宿主代谢的影响，并在AOM/DSS小鼠模型中评估其是否会加重结肠炎相关结肠癌。旨在阐明sennoside A是否破坏结肠上皮屏障的完整性促进结肠炎及结肠癌的发生发展？为明确大黄蒽醌类化合物可能具有的促炎致癌风险的潜在毒性物质基础的确定提供一定的依据。方法利用免疫荧光对相关紧密连接蛋白的检测以及小动物活体成像评价结肠屏障的完整性，并通过16srDNA扩增子测序分析senno⁃side A长期处理对小鼠肠道微生物组成的影响，进一步利用非靶代谢组学分sennoside A对宿主代谢的影响。进一步利用靶向短链脂肪酸代谢组学分析其对短链脂肪酸的影响。Luminex检测sennoside A对诱导结肠组织促炎信号通路的影响。结果从肠黏膜屏障完整性角度出发，评价了sennoside A对于结肠黏膜屏障的两大物理屏障——机械屏障和黏液屏障进行观察。重点检测并分析了sennoside A对肠道微生物菌群结构的影响，发现sennoside A长期处理组的生物标志物，即为sennoside A促进其优势生长的菌群——Akkermansia muciniphila（A.muciniphila），且是一个动态增加的过程。体外菌群培养以确定sennoside A原形，sennoside A微生物代谢混合物以及主要代谢产物Rhein对于A.muciniphila的生长均无显著影响，说明sennoside A对A.muciniphila的生长优势的促进并不是直接作用。进一步分析发现sennoside A对于几种产短链脂肪酸菌属（Coprcoccus，Prevotella，Oscillospira，Ruminococcus）的抑制作用。并对2个主要的产丁酸菌（Clostridium tyrob⁃utyrate和Clostridium butyricum）sennoside A的直接抑菌作用进行考察，说明肠道内的sennoside A可直接抑制产丁酸菌的生长，但不是通过sennoside A原形发挥抑制作用，并确证了sennoside A可显著抑制其生长的菌群及其所引起结肠病理学改变。基于以上，进一步通过粪菌移植实验确证sennoside A所造成肠黏膜屏障的损伤过程，肠道微生物的结构紊乱扮演着不容忽视的角色。senniside A能够引起机体代谢的异常，促进结肠腺瘤的形成。且伴随着STAT3信号通路以及β-catenin信号通路更为强烈的激活，提示着可能产生的恶性转变。结论确证了大黄结合型蒽醌代表性化合物—sennoside A可通过破坏肠黏膜屏障完整性引起结肠长期低水平炎症且促进结肠癌的发展。发现sennoside A可促进A.muciniphila的优势生长以及产丁酸菌群的生长抑制，明确了sennoside A损伤肠黏膜屏障完整性与其所引起的黏液降解菌——外源性纤维素降解菌的平衡破坏密切相关，为大黄蒽醌类化合物使用规避促炎致癌风险提供坚实的实验依据。

关键词：大黄蒽醌类；肠道微生物；肠黏膜屏障；结肠癌

基金项目：国家自然科学基金（81973734；82004124）；南京中医药大学中药一级学科开放项目（2020YLXK20）

通讯作者：陆 茵，E-mail：luyingreen@126.com；韦忠红，E-mail：wzh1225@njucm.edu.cn

番茄红素激活Keap1-Nrf2信号通路抑制诱导型小鼠皮肤癌发生发展

吴媛媛，余苏云，韦忠红，李晓曼，赵 杨，陈文星，王爱云，陆 茵

（南京中医药大学药学院，江苏省中药药效与安全性评价重点实验室，江苏 南京 210023）

摘要：目的阐明目前已知抗氧化活性最强大的药食同源天然产物之一的番茄红素对皮肤癌的化学预防作用及其作用机制。方法建立二甲苯并蒽（DMBA）及12-O-十四烷基-佛波酯-13-乙酸酯（TPA）化学诱导皮肤癌小鼠模型，体内检测番茄红素对皮肤癌发生发展的作用。综合利用药效团模型、分子对接模型、网络拓扑学分析、基因富集分析、基因芯片数据分析、蛋白质间相互作用分析等生物信息学手段，分析番茄红素预防皮肤癌发生发展的关键靶标，挖掘潜在的分子机制。在上皮细胞模型JB6P+细胞里，通过实时荧光定量PCR、Western印迹、免疫组化和免疫荧光等方法，体外检测番茄红素对Nrf2信号通路的影响，进行药效学评价。并利用RNA干扰技术，对Nrf2和p62进行敲减，验证其在番茄红素干预皮肤癌病变过程中的关键作用。结果每天外用涂抹番茄红素8 μmol显著降低DMBA/TPA诱变皮肤癌小鼠模型的发生率及肿瘤的体积。其作用机制是通过上调自噬蛋白p62，诱导Keap1降解,激活转录因子Nrf2，促进下游抗氧化酶系统SOD，CAT，GPx和GR的表达，从而干预DMBA/TPA对上皮细胞氧化应激失衡造成的增殖。结论番茄红素在体内、体外对皮肤肿瘤发生发展的促进阶段有显著的化学预防效应。

关键词：番茄红素；皮肤癌；Keap1；Nrf2；氧化应激

通讯作者：陆 茵，E-mail：luyingreen@njucm.edu.cn

基于肝肠轴探讨异甘草酸镁保护甲氨蝶呤所致肝肠损伤的机制

夏雅雯1，2，钱 程1，2，韩鸿宽1，2，韦忠红1，2，赵 杨1，2，李晓曼1，2，吴媛媛1，2，王爱云1，2，陈文星1，2，陆 茵1，2，3

（1.南京中医药大学药学院，江苏 南京 210023；2.江苏省中药药效与安全性评价重点实验室，江苏 南京 210023；3.江苏省中医药防治肿瘤协同创新中心，江苏 南京 210023）

摘要：目的基于肝肠轴理论探究甲氨蝶呤（MTX）诱导肝肠损伤的内在联系，明确异甘草酸镁（MgIG）对甲氨蝶呤所致肝肠损伤的保护作用并探讨其机制。方法①通过HE染色，GPT和GOT检测及流式细胞术等评价MgIG对MTX诱导的C57BL/6小鼠肝损伤及炎症的保护作用；采用小动物活体成像、免疫荧光染色、短链脂肪酸检测、luminex液相芯片法明确MgIG对MTX所致肠道炎症及渗漏增加的改善作用；②通过在第10，20和30天处理动物，对其肝肠损伤进行评分，探究甲氨蝶呤诱导肝肠损伤发生的先后顺序；③采用荧光原位杂交、CFU菌落计数、LPS检测，结合肝免疫荧光染色验证细菌易位在甲氨蝶呤诱导肝损伤中起到重要作用；④利用16s RNA测序分析寻找MTX处理及给予MgIG治疗前后小鼠肠道中差异菌群，并通过定量PCR对目标菌群的变化进行验证；⑤对目标菌群体外培养并灌胃给予MTX模型小鼠，评价其对肠道屏障及肝损伤的影响；⑥粪菌移植实验阐明肠道菌群在MTX诱导肝肠损伤及MgIG发挥其保护作用过程中是否确实具有重要作用。结果①与对照组相比，ip给予MTX能够引起肝炎症细胞聚集、GPT及GOT水平升高，肠道炎症及通透性增加；给予MgIG治疗显著改善肝肠损伤；②给药10 d后MTX诱导明显的肠道损伤，而肝损伤发生于肠损之后；③ MTX给药过程中，细菌易位至肠道固有层并通过肝肠循环在肝中定植，引起肝LPS水平上升；④与对照组相比，MTX造模组Muribaculaceae丰度升高，乳酸杆菌丰度降低，而这一变化能够被MgIG所逆转；⑤对MTX模型小鼠ig给予乳酸杆菌治疗，小鼠肠道渗漏及肝炎症均被显著改善；⑥MTX组小鼠粪菌移植使正常小鼠出现肠道渗漏及肝炎症，而经MgIG治疗的小鼠粪菌移植至正常小鼠体内并未引起肝肠损伤。结论异甘草酸镁通过恢复乳酸杆菌丰度、降低Muribaculaceae丰度保护肠道屏障，从而抑制MTX引起的肠道菌易位而发挥对肝肠的保护作用。

关键词：肝肠轴；异甘草酸镁；肝肠损伤；甲氨蝶呤

基金项目：江苏省中医药领军人才计划（SLJ0229）；南京中医药大学中药一级学科开放项目（2020YLXK20）

通讯作者：陆 茵，E-mail：luyingreen@126.com

黄芪甘草汤调控SIRT1/FOXO1通路抑制三氧化二砷诱导心肌细胞凋亡的作用机制

徐 兰，苏 敬，刘小凤，董劲曲，侯雯倩，海 洋，刘东玲

（甘肃中医药大学，甘肃 兰州 730000）

摘要：目的探讨SIRT1/FOXO1通路在三氧化二砷（ATO）诱导的心脏毒性机制，以及黄芪甘草汤的防护作用。方法以ATO 5 mg·L-1诱导H9c2细胞损伤模型，采用MTT法、流式细胞术测定不同浓度黄芪甘草汤对心肌细胞存活率、凋亡率以及ROS的影响；Western印迹法测定细胞中SIRT1，FOXO1，Akt和P53的蛋白表达。结果不同浓度的ATO作用于H9c2细胞24 h后，细胞存活抑制率呈浓度依赖性。ATO处理后心肌细胞凋亡率和ROS水平显著升高，SIRT1显著降低，FOXO1水平显著升高（P＜0.05），而加入黄芪甘草汤后均发生逆转。ATO增强EX257对SIRT1的抑制作用，说明ATO引起的心脏毒性部分是通过SIRT1介导。结论ATO引起的心脏毒性与SIRT1密切相关，并且黄芪甘草汤可防治ATO引起的心肌细胞氧化应激与凋亡，该过程可能与SIRT1/FOXO1通路密切相关，为黄芪甘草汤联合ATO用药的开发及临床应用提供理论依据。

关键词：黄芪甘草汤；三氧化二砷；心肌细胞

通讯作者：海 洋，E-mail：cathyhaiyang@163.com

苦豆碱激活胆碱能通路发挥抗结肠炎作用

许霓珊，曹惠慧，卢子滨，余林中，刘俊珊

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的利用小鼠结肠炎模型与Caco-2细胞屏障损伤模型评价苦豆碱的抗结肠炎作用及其机制。方法采用3%葡聚糖硫酸诱导小鼠结肠炎模型，采用脂多糖诱导Caco-2细胞屏障损伤模型；记录小鼠症状并进行疾病活动度指数（DAI）评分；通过HE染色检测结肠组织形态学变化并进行组织病理学评分；通过免疫组化，免疫荧光及Western印迹法检测结肠组织与Caco-2细胞occludin，ZO-1，claudin-1和E-cadherin的表达和分布；通过ELISA检测结肠组织与Caco-2细胞炎症因子白细胞介素6（IL-6）和肿瘤坏死因子α（TNF-α）的表达；通过Western印迹法检测结肠组织与Caco-2细胞中α烟碱型乙酰胆碱受体（α7nAChR）的表达。结果苦豆碱能减缓结肠炎小鼠的体重下降，改善其腹泻、便血、结肠缩短和上皮损伤等症状，提高DAI和组织病理学评分；恢复结肠组织及Caco-2细胞中连接蛋白occludin，ZO-1，claudin-1和E-cadherin的表达及分布；同时降低结肠组织及Caco-2细胞TNF-α和IL-6的表达水平；此外，苦豆碱剂量依赖性促进结肠组织与Caco-2细胞中α7nAChR的活化。结论苦豆碱可能通过激活胆碱能通路发挥抗结肠炎作用。

关键词：苦豆碱；结肠炎；Caco-2

通讯作者：刘俊珊，E-mail：liujunshanty@163.com

基于STAT3/NLRP3双信号通路协同调节髓源抑制性细胞探讨双参抑制肿瘤转移机制

杨葛俊，陈文星

（南京中医药大学药学院，江苏省中药药效与安全性评价重点实验室，江苏 南京 210023）

摘要：目的在肿瘤转移过程中，转移前微环境（PMN）的形成，对于形成肿瘤远端转移灶至关重要。PMN是以免疫抑制为特征，其中免疫负性调控细胞——髓源抑制性细胞（MDSC）发挥重要作用。MDSC可以诱导PMN形成并促进循环肿瘤细胞向PMN迁移，最终导致转移灶形成。基于课题组前期研究表明，丹参作为活血化瘀之要药，其中多种化合物具有改善微循环，提高免疫功能，抗肿瘤作用。人参可通过调节机体免疫微环境，进而发挥抗肿瘤作用。然而，丹参与人参联用（双参）是否能够达到协同抗肿瘤转移的作用，尚未可知。本研究以益气活血中医治则为理论基础，探讨双参是否能够协同作用于MDSC，抑制PMN形成，将有助于达到“已病防变”的治疗目的。方法①体外培养B16F10黑色素瘤细胞，分别设置双参不同剂量组，进行细胞划痕实验、Transwell实验及3D侵袭实验，观察丹参、人参不同剂量配比对B16F10迁移及侵袭能力的影响，其结果利用多药联用效应计算软件SynergyFinder进行协同评分。②通过动物实验探究双参是否能够通过调节免疫微环境有效抑制B16F10肺转移。建立B16F10-luci尾静脉血行转移模型，设置空白对照组、模型组、顺铂组、双参低中高剂量组，连续给药3周后，分别通过小动物活体成像、HE病理染色等技术评价双参对B16F10肺转移的影响。通过流式细胞术及免疫组化分别考察药物对小鼠体内CD4、CD8+T和MDSCs的表达的影响。③探究丹参和人参是否分别通过IL-6/STAT3及NLRP3/胱天蛋白酶1/IL-1β双信号通路影响MDSC在肺部转移灶的募集，进而抑制PMN形成，有效抑制肿瘤转移。分别通过Western印迹法、q-PCR、免疫组化及免疫荧光等方法对上述生物过程和信号通路进行分析。结果①根据B16F10体外迁移侵袭实验的结果，确定丹参-人参（2∶1）联合给药具有协同效应，且协同作用最强。②动物实验结果表明，双参能有效抑制B16F10肺转移，且能够提高小鼠外周血及脾中CD4、CD8+T细胞的表达，减少MDSC的表达。肺组织免疫组化结果显示，双参能够增加肺部转移灶中CD4和CD8+T细胞的浸润，增强荷瘤小鼠免疫功能。③Western印迹法、q-PCR、免疫组化及免疫荧光等实验结果显示，丹参能够抑制IL-6和STAT3在肺部转移灶的表达。人参可以有效抑制NLRP3、胱天蛋白酶1和IL-1β在肺部转移灶的表达。免疫荧光双染结果显示，STAT3及NLRP3与Gr-1存在荧光共定位现象，表明丹参及人参可以分别通过IL-6/STAT3及NLRP3/胱天蛋白酶1/IL-1β双信号通路协同影响MDSC。结论从细胞层面验证了丹参及人参联合用药可协同抑制B16F10黑色素瘤细胞迁移、侵袭；从动物层面验证了丹参及人参联合用药相较于单给药，更能有效抑制B16F10肺转移；从机制方面发现丹参及人参可以分别作用于IL-6/STAT3及NLRP3/胱天蛋白酶1/IL-1β，协同抑制MDSC在肺部转移灶的聚集，最终破坏PMN的形成，达到有效抑制黑色素瘤转移的作用。

关键词：髓源抑制性细胞；丹参；人参；B16F10黑色素瘤细胞

通讯作者：陈文星，E-mail：chenwx@njucm.edu.cn

基于TRPC5探讨山奈酚抗胃癌转移的作用及机制

余苏云1，邓 蕊2，王爱云2，赵 杨1，陆 茵2

（南京中医药大学1.医学院整合医学学院，2.药学院，江苏 南京 210023）

摘要：目的基于细胞运动性探讨天然小分子化合物山奈酚对胃癌转移的药物效应及作用机制。方法首先，通过KM plotter数据库、TCGA等数据库检索分析TRPC5与胃癌发生发展的相关性。并通过IHC、Western印迹法、实时PCR、免疫荧光等验证临床胃癌患者及胃癌细胞中的TRPV4表达水平。随后，通过CRISPR/Cas9构建TRPC5敲除的胃癌细胞，体内外实验验证TRPC5对于胃癌转移的影响。进一步通过化合物结构相似性、CETSA、DARTS、钙成像等实验证明山奈酚能够和TRPC5相结合并能够发挥抑制通道开放的功能。最后，通过免疫荧光、Western印迹法、免疫组化等实验探究山奈酚、TRPC5抑制剂对胃癌细胞运动性相关信号轴的影响。结果通过数据库分析及实验验证表明，胃癌中TRPC5高表达，且其表达水平与之介导的钙内流呈正相关。抑制TRPC5的表达与功能后，胃癌细胞的转移能力显著减弱。化合物结构相似性结果显示山奈酚的结构和TRPC5阳性抑制剂结构相似，且CETSA、DARTS实验表明山奈酚能够与TRPC5直接结合，且体内外结果均表明山奈酚可以抑制胃癌转移。在TIRP数据库探索中发现，TRPC5与细胞运动性及丝状伪足的形成密切相关，而抑制TRPC5的表达与功能均可显著抑制胃癌细胞丝状伪足的形成。进一步的机制探讨发现，TRPC5可以通过p-MLC/ATP/p-cortactin信号轴调控丝状伪足的形成，从而影响胃癌细胞的运动性及其转移进程。结论该研究结合生物信息学分析、临床样本及体内外多项实验，从多个角度确证了TRPC5的表达与胃癌发生发展呈负相关。进一步进行机制探讨，发现TRPC5能够通过p-MLC/ATP/p-cortactin信号轴调控胃癌丝状伪足的形成，揭示丝状伪足在促进胃癌转移中的具体作用机制。发掘存在于多种传统抗肿瘤中药中的有效成分山奈酚，并验证其抗胃癌转移的效应及机制，为临床抗胃癌转移药物的开发提供依据。

关键词：山奈酚；TRPC5；肿瘤转移；细胞运动；丝状伪足

基金项目：国家自然科学基金（81973587；81973734；82003991）

通讯作者：陆 茵，E-mail：luyingreen@126.com

姜黄活性代谢产物四氢姜黄素对乳腺癌作用的hormesis效应规律

曾安琪1，2，戴 瑛1，华 桦1，赵军宁1，2

（1.四川省中医药科学院转化药理与临床应用研究所，四川省中医药转化医学中心，国家中医药管理局中药质量生物评价重点研究室，中医药转化医学四川省重点实验室，四川省道地药材形成原理与品质评价工程研究中心，四川省道地药材系统开发工程技术研究中心，四川 成都 610041；2.四川大学华西药学院，四川 成都 610041）

摘要：目的研究中药姜黄活性代谢产物四氢姜黄素（THC）在不同剂量下对乳腺癌的调治作用。方法利用鼠源4T1细胞对Balb/c小鼠进行接种，建立原位乳腺癌接种模型。接种1周后，将小鼠随机分为模型对照组、空白对照组、THC低剂量组（低1，低2，低3）和高剂量组（高1，高2，高3）共8组，每组10只动物。连续35 d ig给予不同浓度的THC。实验期间每3 d测量小鼠的体重及肿瘤体积。每7 d采血进行流式分析，观察各组小鼠肿瘤免疫微环境中MDSC表型变化及CD4+和CD8+T细胞表型的变化。实验结束后观测各组小鼠体重、生存率、不同级别肿瘤数量、肿瘤大小、脾体积和重量的变化情况。利用ELISA检测肿瘤免疫微环境中细胞因子IFN-γ、TNF-α和IL-2的分泌水平。利用Western印迹法技术和免疫组织化学技术检测样本组织内凋亡、转移相关蛋白的表达变化情况。提取样本组织RNA，利用实时PCR技术检测样本组织内相关基因的mRNA表达变化。以及利用HE染色技术检测各组小鼠重要器官（心、肝、脾、肺、肾）的病理学变化。结果与模型对照组相比，各组小鼠体重及重要器官均未发生显著异常。给予THC低剂量的3个实验组，荷瘤小鼠的肿瘤体积没有发生下降，给予THC第18天后，低3组的荷瘤小鼠肿瘤体积反而较模型组增加，且具有统计学差异（P＜0.05）；而给予THC高剂量的3个组中有2个组（即高2和高3组）荷瘤小鼠的肿瘤体积分别在给予THC 18和15 d之后与模型对照组相比发生了下降，且具有统计学差异（P＜0.05）。Western印迹法、免疫组化和PCR实验结果显示，与模型对照组相比，低2和低3组中Ki-67和MMP-2蛋白的表达增高（P＜0.05）。而高2和高3组中Bcl-2和MMP-2蛋白表达下降（P＜0.05），活化胱天蛋白酶3和TIMP2蛋白的表达增高（P＜0.05）。同时，各组小鼠血液中MDSC以及CD4+和CD8+T表型的变化中，与模型对照组相比，给药第15天起，低2组、低3组以及高3组的小鼠血液中MDSC细胞比例发生下降（P＜0.05）。而与模型对照组相比，给药第21天起低2组、低3组以及高3组的小鼠血液中CD4+和CD8+细胞比例上升（P＜0.05）。ELISA实验检测发现，与模型对照组相比，高2和高3组小鼠血液中IFN-γ、TNF-α和IL-2的含量升高（P＜0.05）；与模型对照组相比，低2和低3组小鼠血液中IFN-γ含量升高（P＜0.05），低2组小鼠血液中IL-2的含量升高（P＜0.05）；低3组小鼠血液中TNF-α含量下降（P＜0.05）。结论THC在乳腺癌的治疗中存在“毒物兴奋效应（hormesis效应）”，低剂量的THC能够促进肿瘤细胞的生长，而高剂量的THC可以抑制肿瘤细胞的生长，调节肿瘤微环境，增强免疫细胞活性。同时发现，一定范围为内低剂量的THC虽然促进肿瘤细胞的生长，但是同时也可以调节肿瘤微环境，增强小鼠免疫功能。

关键词：乳腺癌；四氢姜黄素；肿瘤微环境；hormesis效应

通讯作者：赵军宁，E-mail：zarmy@189.cn

中药单体对细菌群体感应系统的影响

曾 利1，蔡 杨1，凌保东1.2

（成都医学院1.药学院，2.结构特异性小分子药物研究四川省高校重点实验室，四川 成都 610500）

摘要：目的群体感应系统是一种细菌间通信交流机制，与细菌多种毒力因子的产生、生物被膜的形成、芽孢的形成等相关联，是一个感染防治的重要靶点，在细菌的感染致病机理调节中起着核心作用。中药资源丰富，价格低廉，安全性高，毒性小，逐渐成为抗细菌感染研究的热点。近年来有报道中药单体及衍生物可以影响群体感应系统，调节相关毒力因子的表达以及抑制生物被膜的形成。比较研究40种中药单体及衍生物对群体感应系统的影响。方法以紫色杆菌CV026作为研究菌株，分别研究40种中药单体对群体感应系统的作用，筛选出具有抑制活性的中药单体，并探讨中药单体联合作用下对群体感应系统的协同抑制效应。①应用微量肉汤法测定40种中药单体对紫色杆菌CV026的体外最低抑菌浓度（MIC）；②通过报告菌平板法和抑制紫色色素产生实验定性与定量检测亚抑菌浓度下中药单体的群体感应抑制活性；③建立中药单体联合用药的棋盘法，测定药物联合作用下对群体感应系统的协同抑制效应。结果①香豆素、根皮素和大麦芽碱等12种中药单体在亚MIC显著抑制紫色杆菌CV026紫色色素的产生，其中以香豆素和肉桂酸效果最佳，香豆素的MIC为512 mg·L-1，在1/2MIC以及1/4MIC浓度时对群体感应活性抑制率分别为84.93%和66.79%，肉桂酸的MIC为1024 mg·L-1，对群体感应活性抑制率在1/2MIC以及1/4MIC分别为78.78%和56.76%；②12种中药单体不同的联合用药方式产生诱导、无关和抑制群体感应活性，其中以肉桂酸+咖啡酸和肉桂酸+杨梅素的组合抑制效果最佳，肉桂酸和咖啡酸对群体感应活性抑制，在1/4+1/4MIC以及1/8+1/8MIC分别为抑制率75.03%和54.36%，甚至在浓度为1/16+1/16MIC时对群体感应系统活性的抑制率为35.69%，而肉桂酸和杨梅素在浓度为1/4+1/4MIC以及1/8+1/8MIC其抑制率时分别为74.72%和61.74%，在浓度为1/16+1/16MIC时依然有较高的抑制活性，抑制率为41.69%。结论香豆素、咖啡酸和大麦芽碱等12种中药单体在亚MIC下具有显著的群体感应抑制活性；中药单体不同的联合用药方式可产生抑制、无关或诱导群体感应系统的效果，其中以肉桂酸+咖啡酸和肉桂酸+杨梅素的组合协同抑制效果最佳。本研究所筛选的联合用药方式在较低浓度时仍然具有良好的群体感应系统协同抑制活性，不仅可以减少药物的量从而降低药物不良反应，而且可以大大提高疗效。

关键词：中药单体；细菌；群体感应系统

基金项目：国家自然科学基金（81373454）

通讯作者：凌保东，E-mail：lingbaodong@cmc.edu.cn

基于网络药理学和实验验证方法研究皂荚治疗高级别浆液性卵巢癌的分子机制

张博然，张甘霖，王笑民

（首都医科大学附属北京中医医院，北京 100010）

摘要：目的揭示皂角刺（Gleditsiae Spina）的抗卵巢癌的机制。卵巢癌在全球每年导致15.2万人死亡，在女性肿瘤死亡中排名第四，尤其是高浆液性卵巢癌（HGSC），更为恶性，临床预后更差。而目前的中药选择清热解毒治疗只能是手段之一，目前对于皂角刺的研究及应用常被忽视。民间草药医师认为皂角刺具有散结托疮作用，往往在肿瘤方面应用取得意外疗效。因此，本研究希望通过现代研究进行验证，拓展皂角刺的应用。方法采用高效液相色谱法（HPLC）和质谱法（MS）对皂荚的主要活性成分进行鉴定，进行ADME筛选。通过网络药理学方法，使用pharmMapper平台筛选皂荚的成分靶标，通过GEO数据库鉴定正常和HGSC组织中差异表达的基因。此后，利用cytoscape 3.7.2软件构建“活性成分-靶点”网络。利用Bio⁃Genet数据库建立蛋白质-蛋白质相互作用网络，挖掘潜在的蛋白质功能。通过京都基因和基因组百科全书（KEGG）分析来分析生物过程和途径。通过分子对接和分子动力学模拟验证了皂角核心成分与HGSC核心靶点的结合能力，并通过细胞毒性实验在体外证明了皂角的治疗效果，应用免疫印迹实验对影响的核心蛋白及通路进行验证。结果通过HPLC及MS方法得到皂角刺的核心成分有木犀草素、染料木素、D-（+）-色氨酸、熊果酸和小檗碱等，影响到HGSC核心靶点为HPSE，PI3KCA，Akt1和CTNNB1，可以看出，皂角刺可以通过影响蛋白聚糖，干预肿瘤的细胞质基质形成，影响上皮间充质转化的形成，并通过降低PI3K/Akt通路的转化，影响肿瘤发展。而其影响细胞质基质的过程，与中医中对其治疗焦膜病的认识，对其祛焦膜瘀浊的看法相一致。结论皂角主要通过下调肝素酶和β-catenin的表达影响肿瘤细胞质基质的组成，并能调节PI3K-Akt通路，整合多靶点、多通路发挥治疗作用，一定程度揭示了皂角刺治疗卵巢癌的内涵，为今后皂角刺治疗卵巢癌提供了理论依据。

关键词：网络药理学；皂角刺；浆液性卵巢癌

基金项目：国家自然基金（81873111；82074189；82174454）

通讯作者：王笑民，E-mail：wangxiaomin_bhtcm@126.com

大黄素通过核受体Nur77诱导肝星状细胞衰老并减轻肝纤维化

张 峰，陈 利，梁宝瑜，王飞虾，李章昊，杨 婷，郑仕中

（南京中医药大学，江苏省中药药效与安全性评价重点实验室，江苏 南京 210023）

摘要：目的中药大黄常见于临床治疗慢性肝病的复方中，其主要功效物质大黄素对肝纤维化病变的影响机制尚不明确。谷氨酰胺代谢对于肝星状细胞（HSC）活化至关重要。诱导HSC衰老可以逆转肝纤维化。本研究从调控细胞衰老的角度揭示大黄素干预肝纤维化的作用与机制。方法人源HSC-LX2细胞体外培养，检测细胞衰老、谷氨酰胺代谢、谷氨酰胺酶（Gls1）的DNA甲基化及核受体Nur77相关信号分子。小鼠肝纤维化模型给予Nur77 shRNA体内敲减Nur77并给予大黄素维生素A脂质体（体内靶向肝HSC）干预，检测相关指标。结果大黄素上调HSC中衰老相关分子SA-β-Gal、p16、p21及Hmga1的表达并阻滞细胞周期于G0/G1期，提示促进HSC衰老。谷氨酰胺代谢检测显示，大黄素上调细胞内谷氨酰胺含量，但是下调谷氨酸、α-酮戊二酸及三羧酸循环代谢物的含量，提示Gls1及谷氨酰胺代谢受到抑制；这一作用与诱导HSC衰老相关。进一步发现，大黄素促进核受体Nur77转入细胞核，敲减Nur77可以抵消大黄素抑制谷氨酰胺代谢及诱导HSC衰老的作用。机制研究发现，大黄素促进Nur77与DNA甲基化酶DNMT3b相互作用，进而上调DNMT3b催化的Gls1启动子甲基化，导致了Gls1表达及谷氨酰胺代谢受到抑制。谷氨酰胺代谢物α-酮戊二酸可以激活ERK磷酸化进而促进Nur77磷酸化，导致Nur77出核及Gls1启动子甲基化下调，形成了ERK与谷氨酰胺代谢之间的正反馈回路（以α-酮戊二酸和Nur77为关键介导分子）；而大黄素通过抑制ERK和Nur77磷酸化阻断上述正反馈回路，促进Gls1启动子甲基化并抑制其表达。体内研究显示，大黄素维生素A脂质体通过靶向HSC显著减轻肝纤维化小鼠的肝损伤并诱导HSC衰老；体内敲减Nur77能够抵消大黄素维生素A脂质体对HSC谷氨酰胺代谢及衰老的作用。结论大黄素通过调控ERK/Nur77促进Gls1启动子甲基化，导致Gls1表达及谷氨酰胺代谢受到抑制，进而诱导HSC衰老发挥抗肝纤维化作用。

关键词：大黄素；代谢重编程；维生素A脂质体；DNA甲基化

通讯作者：郑仕中，E-mail：nytws@163.com

以荭草苷为关键成分的组合药物设计及其抗心肌缺氧/复氧损伤机制

张明烁，王均琪，万思琦，李志勇，黄秀兰

（中央民族大学，北京 100081）

摘要：目的随着现代中药的发展，以“物质基础基本清楚”、“作用机制基本清楚”为原则的组分中药研宄成为新的热点，初步建立了“标准组分、组效关系、组分配伍、优化设计”的组分中药研发模式。本研宄以荭草苷为关键成分进行组分设计，探寻“试验设计-建立模型-多目标优化”的组分优化方法，探究“以荭草苷为关键成分的组合药物比单个成分在心肌缺氧/复氧损伤（MIRI）模型上的保护作用更好，且组合药物抗MIRI作用机制与调控自噬相关”的假说，为组分优化研究提供方法学上的参考及实验依据。方法研究前期通过筛选条件：①黄酮类成分；②与荭草苷共同存在在同一植物中；③有抗氧化、抗炎等相关药理作用，选出槲皮苷、牡荆素、槲皮素、木犀草素、异荭草素5个成分，采用CCK-8法检测各成分对H9c2细胞存活率的影响，采用三气培养箱混合气体法建立心肌细胞缺氧/复氧损伤模型，通过CCK-8法和LDH法研究各成分对H9c2缺氧/复氧损伤细胞存活率、损伤率的影响。通过均匀设计得到荭草苷、槲皮苷、牡荆素的不同配比组合，利用CCK-8法和LDH法检测各配比对H9c2缺氧/复氧损伤模型细胞存活率和损伤率的影响；检测培养基中NO含量，利用R语言建立逐步回归模型，基于模拟退火算法理论，用Python编程解出当综合药效最佳时各成分的给药浓度，即最佳配比组合OQV-e，对OQV-e进行药效学实验验证，通过Western印迹法检测H9c2细胞内LC3，Beclin-1，p-JNK和JNK的蛋白表达水平，评价最佳配比对缺氧/复氧损伤H9c2细胞内JNK通路及细胞自噬的影响。结果① 荭草苷（0～200 μmol·L-1），槲皮苷（0～200 μmol·L-1），牡荆素（0～80 μmol·L-1），异荭草苷（0～200 μmol·L-1）孵育36 h内对H9c2细胞增殖无明显影响；除异荭草苷外，荭草苷、槲皮苷、牡荆素均能显著性下调细胞损伤率。②OQV-e能显著提高H9c2心肌细胞存活率，降低细胞损伤率，降低NO含量，且OQV-e比其他单个成分降低细胞损伤率效果更明显（P＜0.05）。③与模型组相比，OQV-e的p-JNK/JNK的比值降低，LC3-Ⅱ/LC3-Ⅰ比值显著性降低，Beclin-1的表达水平明显升高（P＜0.05）。结论①结合文献调研与初步药效实验结果，选择荭草苷、槲皮苷、牡荆素进行组合药物设计；②采用均匀设计为试验设计方法，以H9c2心肌细胞缺氧/复氧损伤为实验研究模型，逐步回归分析为建模准则，模拟退火算法为计算理论，计算出最佳配比组合OQV-e，即荭草苷12.55 μmol·L-1，槲皮苷 39.99 μmol·L-1，牡荆素19.99 μmol·L-1。③OQV-e抗MIRI可能是通过抑制JNK信号通路，抑制过度自噬，进而达到保护缺氧/复氧损伤心肌的目的。

关键词：荭草苷；组合药物设计；缺氧/复氧损伤

基于网络药理学与分子对接研究大花红景天防治炎症风暴作用机制及实验验证

赵万花，王平义，宋 丹，顾 霞，田 雨，李文华

（西藏民族大学医学部，陕西 咸阳 712082）

摘要：目的通过网络药理学与分子对接方法探究藏药大花红景天防治炎症风暴的的潜在生物学机制，并通过动物实验初步验证。方法从化学专业数据库中收集大花红景天的有效成分和相关靶点；通过GeneCards、OMIM数据库收集细胞因子风暴相关基因；应用Cytoscape3.7.2软件构建大花红景天活性成分-细胞因子风暴靶点调控网络图；利用STRING数据库，构建蛋白质-蛋白质相互作用网络（PPI）。采用R软件及DAVID数据库进行GO富集分析和KEGG富集分析。运用autoDock软件进行分子对接。通过动物实验对IL-4信号通路进行了验证。结果经过筛选和去除重复值后，共获得与防治细胞因子风暴相关的16种活性成分和49个有效靶点。PPI网络表明，IL-6，STAT3，SRC，Akt1，EGFR和CXCL8等是关键的靶蛋白。GO分析发现大花红景天在抗细胞因子风暴时主要涉及调控细胞凋亡、基因表达、细胞因子活性、免疫应答等过程。KEGG通路分析发现其主要涉及TNF，PI3K-Akt和kaposi sarcoma-associated herpesvirus infection信号通路等核心靶点。分子对接结果显示，山奈酚和红景天苷与SARS-CoV-23CL水解酶和ACE2均具有较好的亲和力。动物实验证实了大花红景天能有效抑制炎症因子风暴并提高ACE2的含量。结论大花红景天能通过与SARS-CoV-23CL水解酶和ACE2结合作用于IL-6等靶点,增加ACE2含量，减轻炎症因子风暴引起的肺损伤。

关键词：大花红景天；细胞因子风暴；新型冠状病毒肺炎；网络药理学；山奈酚；红景天苷；分子对接

通讯作者：李文华，E-mail：xzmylwh3311779@163.com

基于CCL20/CCR6探讨人参皂苷Rh1调控乳腺癌细胞外渗的作用机制

郑维维，张 珊，唐 彧，王飞辉，周玥珂，邹得方，陆 茵，王爱云

（南京中医药大学药学院，江苏省中药药效与安全性评价重点实验室，江苏省中医药防治肿瘤协同创新中心，江苏 南京 210023）

摘要：目的研究人参皂苷Rh1调控肿瘤细胞外渗的机制。方法首先借助微流控芯片模拟肿瘤转移过程，借助斑马鱼模型进行验证；并结合转录组学测序的方法对其机制进行探讨，最后通过质粒转染、siRNA干扰技术对该机制进行验证。结果①微流控芯片结果显示Rh1显著抑制肿瘤细胞外渗过程，斑马鱼实验验证这一结果；②体外肿瘤细胞-内皮细胞共培养后，对肿瘤细胞的转录组测序结果显示Rh1可能通过抑制CCL20的表达发挥作用；通过质粒转染肿瘤细胞过表达CCL20后，借助斑马鱼模型发现，肿瘤细胞出血管能力增强；③借助一系列体外侵袭实验和层位移实验发现，过表达CCL20的肿瘤细胞表现出更强的侵袭性和破坏内皮层的能力。结论肿瘤细胞外渗过程中分泌的CCL20，一是通过自分泌增强自身的侵袭性，二是通过旁分泌作用于内皮层，破坏内皮层的紧密连接，而Rh1通过抑制肿瘤细胞CCL20的表达，逆转这一过程。

关键词：人参皂苷Rh1；肿瘤细胞；细胞外渗；CCL20/CCR6

通讯作者：王爱云，E-mail：way9815@163.com

黄芩苷通过抑制星形胶质细胞肿胀并抑制TRPV4和AQP4的通道开放治疗大鼠脑缺血再灌注所致脑水肿

郑晓宇，宋文婷，张业昊，曹 慧，刘建勋

（中国中医科学院西苑医院基础医学研究所，中药药理北京市重点实验室，北京 100091）

摘要：目的研究黄芩苷对脑缺血再灌注损伤所引起的脑水肿大鼠的药效学作用及作用机制。脑缺血后，星形胶质细胞表现为增生、肿胀并形成瘢痕，瞬时受体电位香草素受体4型通道蛋白（TRPV4）为钙离子通道，水通道蛋白4（AQP4）为水通道，2种通道蛋白在星形胶质细胞膜上均有较高表达，TRPV4和AQP4的开放和关闭与星形胶质细胞肿胀密切相关。方法采用大脑中动脉闭塞法（线栓法）制作SD大鼠脑缺血模型，并于缺血1.5 h后拔出线栓实现再灌注，将大鼠随机分为正常组、模型组、黄芩苷10和20 mg·kg-1组，于大鼠脑缺血时进行尾静脉注射给药，缺血24 h后取材，分别测定大鼠脑缺血后6和24 h的神经行为学、脑含水量、脑梗死面积，Western印迹测定脑组织中GFAP，TRPV4和AQP4的含量，脑片病理切片后采用苏木素-伊红染色、免疫组织化学染色观察病理指标。建立SVGp12星形胶质细胞氧糖剥夺（OGD）损伤模型，观察黄芩苷对OGD损伤后SVGp12细胞活力的影响。结果与正常组相比，模型组大鼠的神经行为学评分、脑含水量和脑梗死面积均显著增加（P＜0.01），GFAP，TRPV4和AQP4的表达均明显增加（P＜0.01）。与模型组相比，黄芩苷10和20 mg·kg-1组均可显著降低大鼠的神经行为学评分、脑含水量和脑梗死面积（P＜0.05），GFAP，TRPV4和AQP4的表达也显著降低（P＜0.05）。病理学可见，正常组大鼠脑组织排列致密且规则，模型组大鼠大脑皮质结构疏松，神经元减少、紧缩并颜色加深、细胞周围间隙增大，星形胶质细胞出现明显的增生与肿胀；与模型组比较，黄芩苷组神经元数量增加，仅有少量神经元紧缩，星形胶质细胞增生与肿胀的情况有明显改善，均可有效改善大鼠脑皮质结构损伤。免疫组织化学染色结果显示，模型组与正常组相比，GFAP，AQP4和TRPV4的表达均增加，黄芩苷20 mg·kg-1组可不同程度降低其表达。细胞实验结果显示，黄芩苷可以显著改善OGD诱导后SVGp12细胞活力（P＜0.01）。结论黄芩苷可有效改善大鼠脑缺血再灌注损伤所致的脑水肿，可能与抑制星形胶质细胞肿胀并抑制TRPV4和AQP4的通道开放等有关。

关键词：黄芩苷；星形胶质细胞；脑缺血再灌注

通讯作者：刘建勋，E-mail：liujx0324@sina.com

白花蛇舌草抗肝癌作用

周春红，许霓珊，曹惠慧，余林中，区锦莹，刘俊珊

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的通过斑马鱼异体肝癌细胞移植模型和Hep3B细胞体外实验研究白花蛇舌草不同极性部位的抗肝癌作用。方法采用浸泡给药的方式确定白花蛇舌草的水、正丁醇、乙酸乙酯和石油醚提取部位对斑马鱼的体内毒性；建立斑马鱼异体荧光标记肝癌细胞移植模型，拍照记录斑马鱼卵黄囊内细胞荧光强度的变化，HE染色观察肝癌细胞的数目，评价白花蛇舌草不同极性部位提取物的体内抗肝癌作用；通过Hoechst染色观察斑马鱼卵黄囊内肝癌细胞的凋亡；通过MTT、Annexin V-FITC/PI双染实验和细胞周期分析检测白花蛇舌草乙酸乙酯部位对Hep3B细胞的细胞毒性、凋亡以及周期的影响；通过Western印迹法检测Hep3B细胞给药后PARP，Nur77，p-Nur77，JNK和p-JNK的表达。结果白花蛇舌草乙酸乙酯部位能显著抑制斑马鱼异体移植肝癌细胞的生长，时间、剂量依赖性地抑制Hep3B细胞增殖，促进其凋亡，并阻滞其细胞周期于G2/M期。此外，该提取物能降低Hep3B细胞中Nur77，p-Nur77，JNK和p-JNK的表达。结论白花蛇舌草乙酸乙酯部位可能通过抑制Nur77/JNK信号通路发挥抗肝癌作用。

关键词：白花蛇舌草；抗肝癌；斑马鱼

通讯作者：刘俊珊，E-mail：liujunshanty@163.com

中药复方制剂对2型糖尿病视网膜病变小鼠的血管保护作用

周 赫1，2，刘 博1，王 鑫1，丁 涛1，温富春1，徐惠波1

（1.吉林省中医药科学院，吉林 长春 130012；2.长春中医药大学，吉林 长春 130117）

摘要：目的观察由黄连黄芪等中药组成的中药复方制剂对2型糖尿病视网膜病变（DR）小鼠的血管保护作用。方法雄性C57BL/6小鼠预留20只作为空白组，普通饲料喂养，其余动物以高糖高脂饲料（kk饲料）喂养加ip给予链脲佐菌素（100 mg·kg-1）建立2型糖尿病小鼠模型，每天分别ig给予羟苯磺酸钙0.3 g·kg-1或中药复方0.36、0.72和1.43 g·kg-1，连续给药10周。测量小鼠给药后血糖，检测糖耐量（AUC）、胰岛素C肽、肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）、IL-6、IL-18、NF-κB、血栓素B2（TXB2）和6酮前列腺素F1α（6-keto-PGF1α）；HE染色观察动物胰腺和视网膜病理变化，并检测视网膜厚度和视网膜神经节细胞（RGC）数；TUNEL染色观察视网膜细胞凋亡情况并计算凋亡指数。免疫组织化学方法检测小鼠视网膜组织中VEGFA，VEGFB，VEGFR2和GFAP的表达。结果与空白对照组相比，小鼠造模后血糖、TNF-α，IL-1β，IL-6，IL-18，NF-κB和TXB2水平均显著升高（P＜0.05），胰岛素C肽和6-keto-PGF1α水平均显著降低（P＜0.01），胰岛及视网膜组织损伤较重，小鼠视网膜厚度明显变薄，RGCs数量明显下降（P＜0.01）。与模型对照组相比，中药复方制剂ig给药能显著降低血糖、TNF-α，IL-1β，IL-6，IL-18，NF-κB和TXB2水平（P＜0.05），升高胰岛素C肽和6-keto-PGF1α水平（P＜0.05）；HE染色结果显示，中药复方制剂给药后小鼠胰腺内可见体积较大、数量较多的胰岛组织散在分布，胰岛数量高于模型对照组；与模型对照组比较，各给药组眼内各结构相关形态变化得到有效改善，视网膜神经节细胞层内的RGC数量明显增多，视网膜厚度明显增加（P＜0.05）。各剂量中药复方组细胞凋亡指数显著降低（P＜0.05）；免疫组化结果显示，各剂量中药复方组不同程度减少视网膜组织中VEGFA，VEGFB，VEGFR2和GFAP的表达量。结论中药复方制剂的应用，证明了该组合物具有降血糖作用和视网膜保护作用，其作用机制可能与保护胰岛功能与形态、改善炎症反应、减少视网膜微血管损伤有关。

关键词：中药；复方；糖尿病视网膜病变；胰岛功能；炎症反应；视网膜微血管

基金项目：国家重大科技专项（2017ZX09301031）；吉林省科技发展计划（20170311012YY）

通讯作者：徐惠波，E-mail：xhb_6505@163.com

丹参酮ⅡA靶向Ang2/Tie2轴诱导缺血性损伤和肿瘤血管正常化

邹 伟1，韦忠红1，王爱云1，2，陆 茵1，2

（南京中医药大学1.江苏省中药药效与安全性评价重点实验室，2.江苏省中医药防治肿瘤协同创新中心，江苏 南京 210023）

摘要：目的丹参酮ⅡA（Tan ⅡA）是丹参的活性成分，在改善血液循环和延缓肿瘤进展方面具有重要作用。然而对Tan ⅡA治疗效果的潜在机制尚不完全了解。本研究建立HT-29人结肠癌异种移植和后肢缺血动物模型，以研究Tan ⅡA在调节异常血管系统中的作用。方法选取BALB/c Nude小鼠，建立异病同体小鼠模型，造模成功后ig给予Tan ⅡA 10，30和90 mg·kg-1，采用激光多普勒血流仪分别对术前、术后以及术后第7，14和21天下肢血流进行监测，通过下肢外观评分、腓肠肌组织H&E染色、免疫组化染色和免疫荧光染色评价Tan ⅡA对缺血下肢血管正常化的影响；通过分析结肠癌皮下移植瘤相对增殖率、肿瘤大小、肿瘤质量和血管渗漏及结合免疫荧光和免疫组化等检测方法评价Tan ⅡA对肿瘤组织血管正常化的影响。结果Tan ⅡA能显著促进缺血损伤后的血流，减轻缺氧，改善肌肉质量，改善病理损伤。Tan-ⅡA还可促进HT-29肿瘤血管结构的完整性，减少血管渗漏，减轻缺氧。此外，Tan ⅡA显著降低了这2种病理状态下Ang2的分泌，并增强Tie2的激活，导致血管通透性降低和血管完整性提高。在机制上，Tan ⅡA通过靶向Ang2-Tie2-Akt-MLCK级联反应维持血管稳定性。结论Tan ⅡA通过调节Ang2/Tie2信号轴使肿瘤和缺血性损伤中的血管正常化。

关键词：丹参酮ⅡA；下肢缺血；结肠癌；血管正常化；Ang2/Tie2轴

基金项目：国家自然科学基金（81973587；81973734；82003991）

通讯作者：陆 茵，E-mail：luyingreen@126.com

论文摘要

14-Deoxygarcinol,a polyisoprenylated benzophenone from Garcinia cambogia,ameliorates inflammatory responses in adipose tissue via suppressing NLRP3 inflammasome

CHEN Jia-li＊,FENG Zhe-ling＊,CHEN Cheng,ZHU Jian-zhong,LIN Li-gen

（State Key Laboratory of Quality Research in Chinese Medicine,Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVEInterleukin（IL）-1β,one of the principal inflammatory cytokines mainly secreted by mono⁃cytes and macrophages,is produced by cleavage of the inactive pro-IL-1β precursor by caspase-1 via the NLRP3 inflam⁃masome complex.The fruits of Garcinia cambogia（Clusiaceae）are widely developed as health products for anti-obese purpose.14-deoxygarcinol（DOG）is a polyisoprenylated benzophenone from the fruits of G.cambogia,which showed potent anti-inflammatory effect in our previous study.The objective of this study was to explore the anti-inflammatory mechanism of DOG and its roles in alleviating adipose tissue inflammation and insulin resistance.METHODSThe antiinflammatory effect of DOG was evaluated on LPS plus nigericin-induced THP-1 macrophages.The expression of NLRP3 inflammasome complex proteins was analyzed by Western blotting,immunofluorescence staining and co-immu⁃noprecipitation.The pro-inflammatory cytokines levels were determined by ELISA kits.RESULTSDOG increased the expression of Sirtuin 2（SIRT2）deacetylase and enhanced its deacetylating activity to suppress the NLRP3 inflamma⁃some activation and IL-1β secretion in THP-1 macrophages.Moreover,DOG attenuated macrophage conditioned mediuminduced inflammatory responses in adipocytes and blocked THP-1 macrophages migration towards 3T3-L1 adipocytes.CONCLUSIONDOG attenuated the inflammatory crosstalk between macrophages and adipocytes through SIRT2-mediated NLRP3 inflammasome inhibition,which might be used for the treatment of adipose tissue inflammation-related metabolic disorders.

Key words：14-deoxygarcinol;Sirtuin 2;interleukin-1β;macrophages;NLRP3 inflammasome

Foundation item：National Natural Science Foundation of China（81872754）;Research Fund of University of Macau（MYRG2018-00037-ICMS）;and Science and Technology Development Fund,Macao SAR（FDCT 0031/2019/A1）

Corresponding author：LIN Li-gen,E-mail：ligenl@um.edu.mo

＊Co-first author.

Inhibition of colorectal cancer by ursolic acid via noncanonical Hedgehog signaling pathway

CHEN Li1,SUN Qiang1,ZENG Sha1,ZHAO Hui1,LIU Mao-lun1,YANG Han1,REN Shan1,MING Tian-qi1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVETo identify the inhibitory effect of ursolic acid on the colorectal cancer HCT116 cells in vitro and in vivo,and to explore the underlying mechanism.METHODSThe smoothened（SMO）gene-silenced human colorectal cancer HCT116hSMO-cell line was established by transfection with the lentivirus carrying SMO shRNA.The cytotoxic effect of ursolic acid on HCT116hSMO-cells was determined by MTT assay.The effect of ursolic acid on the migration of HCT116hSMO-cells was studied by wound healing assay.The effect of ursolic acid on apoptosis of HCT116hSMO-cells was explored by Hoechst33342/PI double staining and flow cytometry.The effects of ursolic acid on the expressions of apoptotic marker gene Bcl-2,Bax,caspase-3 and caspase-9 were measured by real-time quantitative RT-PCR（RT-qPCR）and Western blotting（WB）analysis.RT-qPCR and WB were used to examine the relationship between GLI1,c-Myc expression and PI3K/Akt pathway to further investigate the mechanism of GLI1 activation in HCT116hSMO-cells.The effects of ursolic acid on the expressions of GLI1,p-Akt,Akt,c-Myc,SHH and SUFU of nonca⁃nonical Hedgehog pathway were evaluated by RT-qPCR and WB assays.Xenograft nude mouse model bearing HCT116hSMO-cells was established and intraperitoneally treated with ursolic acid to investigate the effect on tumor growth in vivo.The body weight and tumor size of mice were assessed regularly every 2 d.The effect of ursolic acid on the apoptosis of tumor tissue was determined by TUNEL assay.The expressions of Bcl-2,Bax,GLI1,p-Akt,Akt,c-Myc,SHH,SUFU mRNA and proteins were measured by RT-qPCR and WB.The levels of Bcl-2,Bax,GLI1,p-Akt,c-Myc and SHH proteins in tumor tissues were also evaluated by immunohistochemistry.RESULTSUrsolic acid significantly inhibited the growth and migration of HCT116hSMO-cells in vitro,compared with the control（P＜0.05）.Meanwhile,ursolic acid also induced apoptosis of HCT116hSMO-cells in vitro（P＜0.05）.Furthermore,SC79（Akt activator）enhanced the expressions of p-Akt,GLI1 and c-Myc,which could be abolished by ursolic acid,and the effect was equal to Akt inhibitor LY294002.The expressions of Bcl-2,GLI1,p-Akt,c-Myc,SHH mRNA and proteins were reduced by ursolic acid,while the levels of Bax and SUFU were increased.Ursolic acid could inhibit the growth and induce the apoptosis of colorectal cancer xeno⁃graft in vivo.Similarly,lower levels of Bcl-2,GLI1,p-Akt,c-Myc and SHH,and higher expression of Bax and SUFU were noted in ursolic acid-treated mice.CONCLUSIONUrsolic acid can inhibit the growth and induce apoptosis of HCT116hSMO-cells both in vitro and in vivo.And the mechanism is related to the suppression of PI3K/Akt-mediated noncanonical Hedgehog signaling pathway.

Key words：ursolic acid;colorectal cancer;noncanonical Hedgehog signaling;apoptosis

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;Excellent Talent Program of Chengdu University of Tra⁃ditional Chinese Medicine of China（YXRC2019002;ZRYY1917）;and Open Research Fund of the State Key Laboratory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Effects of polygonatum polysaccharide on p38 MAPK/N-cadherin in zebrafish with Alzheimer disease

CHEN Yi-fei1,LIU Kai-fei1,WU Shi-min1,LI Xun-yi1,HUANG Yao1,HUANG Dian-feng1,JIANG Cai-wu2

（1.Guilin Medical University,Guilin 541199,China;2.Guangxi University of Chinese Medicine,Nanning 530200,China）

Abstract：OBJECTIVETo study the effect of polygonatum polysaccharide on zebrafish with Alzheimer disease.METHODSZebrafish were trained in T maze for 7 d.The 40 zebrafish successfully trained were divided into 4 groups：blank group,model group,positive group and polygonatum polysaccharide group.Model group,positive group and polygonatum polysaccharide group were put in AlCl3100 μg·L-1for 6 d.The positive group was exposed to Huperzine A solution 4 μg·L-1,and the polygonatum polysaccharide group was exposed to polygonatum polysaccharide solution 6 g·L-1for 6 d.The model group was not treated,and the blank group was not treated.Each stage of zebrafish was recorded by video,and the time of each group in the EC region was analyzed.After administration,the brain tissue was taken out and the expression of N-cadherin,P38 and p-P38 protein factors was determined by Western blotting.RESULTSIn behavior,the analysis of the time spent in the EC area,the blank group,the positive group and the polygonatum polysac⁃charide group were compared with the model group,respectively,there were statistically significant differences（P＜0.05）.At the protein level,compared with the model group,the P38 and p-P38 proteins in the positive group and the polygonatum polysaccharide group were down-regulated,while the N-cadherin protein was up-regulated,with statistical difference（P＜0.05）.CONCLUSIONPolygonatum polysaccharide can improve the learning and memory ability of zebrafish with Alzheimer disease by up regulating the protein level of N-cadherin and hindering P38 phosphorylation.

Key words：Alzheimer disease;zebrafish;polygonatum polysaccharide;N-cadherin;p38 MAPK

Myricetin inhibits interferon- γ-induced programmed death ligand-1 and indoleamine 2,3-dioxygenase 1 expression in lung cancer cells

CHEN Yu-chi1,HE Xin-ling1,QI Lu1,SHI Wei1,YUAN Luo-wei1,HUANG Mu-yang1,XU Yu-lian1,CHEN Xiu-ping1,ZHANG Le-le1,LU Jin-jian1,2

（1.State Key Laboratory of Quality Research in Chinese Medicine,Institute of Chinese Medical Sciences,University of Macau,Macao,China;2.MoE Frontiers Science Center for Precision Oncology,University of Macau,Macao,China）

Abstract：OBJECTIVEProgrammed death ligand-1（PD-L1）and indoleamine 2,3-dioxygenase 1（IDO1）are immune checkpoints which can be induced by interferon-γ （IFN-γ）in the tumor microenvironment,leading to immune escape of tumors.Myricetin（MY）is a flavonoid distributed in many edible and medicinal plants.The aim of this study is to clarify the effect and the mechanism of MY on inhibiting IFN-γ-induced PD-L1 and IDO1 in lung cancer cells.METHODSExpressions of PD-L1 and major histocompatibility complex-I（MHC-I）were evaluated by flow cytometry and Western blotting,and the expression of IDO1 was measured by Western blotting.qRT-PCR was used to detect their mRNA levels.The function of T cells was evaluated using a co-culture system consist of lung cancer cells and the Jurkat-PD-1 T cell line that overexpressing PD-1.Molecular docking analysis,Western blotting and immunofluorescence were used for mechanism study.RESULTSMY potently inhibited IFN-γ-induced PD-L1 and IDO1 expression in human lung cancer cells,while didn't show obvious effect on the expression of MHC-I.In addition,MY restored the survival,proliferation,CD69 expression and interleukin-2（IL-2）secretion of Jurkat-PD-1 T cells suppressed by IFN-γ-treated lung cancer cells in the co-culture system.Mechanistically,IFN-γ up-regulated PD-L1 and IDO1 at the transcriptional level through the JAK-STAT-IRF1 axis,which was targeted and inhibited by MY.CONCLUSIONOur research revealed a new insight into the anti-tumor effects of MY which inhibited IFN-γ-induced PD-L1 and IDO1 expression,supporting the potential of MY in anti-tumor immunotherapy.

Key words：programmed death ligand-1;indoleamine 2,3-dioxygenase 1;myricetin;interferon-γ;lung cancer

Foundation item：Science and Technology Development Fund,Macau SAR（0129/2019/A3;176/2017/A3）;and University of Macau（MYRG2018-00165-ICMS）

Corresponding author：LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Pharmacological effects of volatileoil from Alpinia officinaruim in treating cardiovascular diseases

CHENG Wen-jing1,JING Yong-shuai1,ZHANG Dan-shen1,ZHENG Yu-guang2,WU Lan-fang2

（1.College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China;2.College of Pharmacology,Hebei University of Chinese Medicine,Shijiazhuang 050200,China）

Abstract：Galangal（Alpinia officinaruim Hance）is the rhizome of the perennial herb belonging to Zingiberaceae family.There are many active components in galangal,such as volatile oil,flavonoids,terpenoids,phenylpropanoids and glycosides,among which the content of volatile oil is higher.The bioactivities of galangal volatile oil on health effect includesanti-inflammatory,anti-hypertension,anti-oxidation and prevention of cardiovascular diseases.Cardiovascular disease（CVD）is a kind of diseases related to circulatory system,which is also called circulatory system diseases.Over the past decade,the number of people dying from CVD has increased by 12.5%worldwide,and it is now the leading cause of human death worldwide.Studies have shown that galangal volatile oil has good pharmacological effects in treating CVD.①Regulation of glucose and lipid metabolism：studies have found that abnormal lipid metabolism can lead to obesity,diabetes,CVD and other diseases.The serum total triglyceride（TG）content in liver and serum will increase in patients with abnormal fat metabolism.The results showed that the volatile oil of galangal could increase the excretion of neutral cholesterol,significantly reduce liver TG and serum TG,and thus regulate glucose and lipid metabo⁃lism,prevent lipid deposition and prevent CVD.②Improving insulin resistance（IR）：inhibition of inflammatory cytokines such as IL-1,IL-6 activation and expression of TNF-α,improves IR,thereby protecting myocardium from IR-mediated damage.Through the establishment of endothelial cell injury model induced by high glucose in vitro,it was found that the volatile oil of galangal can significantly reduce the secretion of pro-inflammatory cytokines TNF-α and IL-8,and inhib⁃it the expression of ICAM-1 and VCAM-1 induced by high glucose,suggesting that it has protective effect on endothelial dysfunction and inflammation induced by high glucose.③Regulate blood oxygenation：during acute myocardial hypoxia,the activity of free radical scavenging system is decreased,and oxygen free radicals are produced in large quantity,which reacts with unsaturated fatty acids on the cell membrane and forms lipid peroxidation,resulting in myocardial structural damage.The results showed that the water extract of Galangal could reduce the content of MDA in blood and protect the SOD activity of ischemic and hypoxic myocardium.④Protective effect of vascular endothelial cells（ES）：ES injury is the pathological basis of some cardiovascular diseases.The results showed that the volatile oil of galangal had a protective effect on ES apoptosis.Compared with the morphology and activity of ES treated with oxidized LDL,galan⁃gal volatile oil could ameliorate these morphological changes and improve cell viability.⑤Antiplatelet agglutination：inhibit platelet aggregation and thromboxane release,improve blood circulation,and have obvious anti-thrombotic effect,which has a good effect on the treatment and prevention of cardiovascular diseases.The results showed that the volatile oil of galangal had inhibitory effect on platelet aggregation and anticoagulant effect.In conclusion,the volatile oil of galangal can be used to prevent and treat cardiovascular diseases.Based on the mechanism of CVD,this study summa⁃rized the role of the essential oil of Alpinia officinaruim in CVD,providing basis for the clinical application of alpiniaoffici⁃nalis essential oil in the prevention and treatment of CVD and the development of new drugs.

Key words：Alpinia officinaruim;essential oil;Cardiovascular disease;pharmacological effect

Foundation item：The S&T Program of Hebei Province（H2021423057）

Corresponding author：WU Lan-fang,E-mail：wulanfang757@163.com

In vivo and in vitro inductively coupled plasma mass spectrometry quantification of minerals and heavy metals in Qishiwei Zhenzhu pill and their effect on intestinal flora

FU Ke1＊,ZHANG De-wei2＊,SONG Ying-lian1＊,XU Min1,WU Rui-xia1,XIONG Xue-qing2,LIU Xian-wu1,WU Lei1,GUO Ya2,ZHOU You1,LI Xiao-li1,WANG Zhang3

（1.College of Pharmacy,3.College of Ethnomedicine,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Wanzhou Institute for Drug and Food Control,Chongqing 404000,China）

Abstract：OBJECTIVECerebral ischemia or ischemic stroke is due to insufficient blood supply to the brain,which causes hypoxia or ischemia in some areas.This work aimed to quantify the minerals and heavy metals in Qishiwei Zhenzhu pill in vivo and in vitro,analyze its effect on the types and abundance of intestinal flora,and study its mechanism on inflammation and apoptosis pathways as a treatment for cerebral ischemia.METHODSMicrowave digestion and induc⁃tively coupled plasma mass spectrometry（ICP-MS）were used to determine the minerals and heavy metals in 10 batches of Qishiwei Zhenzhu pill in vitro.With the use of the middle cerebral artery obstruction（MCAO）model,ICP-MS was applied to determine the content of minerals and heavy metals in hepatic portal vein blood,abdominal aortic blood,brain,liver,kidney,hair,urine and feces at different time periods.On this model,the ileum,cecum,and colon tissues were tested for intestinal pathology,and 16S rRNA was used for sequencing.Species taxonomy,α diversity,and spe⁃cies microbial composition and structure analysis were also performed.Polymerase chain reaction and Western blotting were employed to determine the mRNA and protein expression of p38 MAPK,caspase-3,IL-1β and TNF-α in the isch⁃emic brain tissues of rats.RESULTSThe average content of heavy metals in the 10 batches of Qishiwei Zhenzhu pill samples is in the descending order Hg>Cu>Pb.Significant differences in the metal elements are found among Qishiwei Zhenzhu pill from different manufacturers but not among the different batches of the same manufacturer.An extremely low content of heavy metals are absorbed into the blood or accumulated in the brain,liver,kidney,and other tissues.Stool is the main excretion route of minerals and heavy metals from Qishiwei Zhenzhu pill.This medicine helps repair the intestinal mucosa in MCAO rats.At the phylum level,it can regulate the abundance of Firmicutes and Proteobacteria in the intestinal flora of rats with cerebral ischemia.At the genus level,it can adjust the abundance of Escherichia Shigella.At the species level,it can adjust the abundance of Lactobacillus yoelii and Lactobacillus reuteri.Cluster classification results show that Qishiwei Zhenzhu pill can improve the intestinal flora of rats with cerebral ischemia,reduce the mRNA and protein expression of caspase-3 and IL-1β in rat brain tissues,and have a tendency to decrease the mRNA expres⁃sion of p38 MAPK and TNF-α.CONCLUSIONQuantifying the minerals and heavy metals in Qishiwei Zhenzhu pill in vivo and in vitro will help improve their quality standards.Minerals and heavy metals are mainly excreted in feces,accumu⁃late in extremely low levels in various tissues,and do not damage the intestinal mucosa.The effective material basis of Qishiwei Zhenzhu pill in treating cerebral ischemia may be related to their Li,Cr,and Cd elements.These pills can improve the environment of intestinal flora,and their mechanism of treatment for cerebral ischemia may be related to the down-regulation of IL-1β inflammatory factor and inhibition of cell apoptosis.

Key words：Qishiwei Zhenzhu pill;cerebral ischemia;inductively coupled plasma mass spectrometry;minerals,heavy metals;intestinal flora

Corresponding author：WANG Zhang,E-mail：wangzhangcqcd@cdutcm.edu.cn

＊Co-first author.

Magnolol attenuates right ventricular hypertrophy and fibrosis in hypoxia-induced pulmonary arterial hypertensive rats through inhibition of the JAK2/STAT3 signaling pathway

FU Min-yi1,LUO Fang-mei2,WANG E-li3,JIANG Yue-ping1,LIU Shao1,LUO Xiu-ju4,PENG Jun3,5,LIU Bin1

（1.Department of Pharmacy,Xiangya Hospital,Central South University,Changsha 410008,China;2.Department of Pharmacy,Hunan Children′s Hospital,Changsha 410007,China;3.Department of Pharmacology,5.Hunan Provincial Key Laboratory of Cardiovascular Research,Xiangya School of Pharmaceutical Sciences,Central South University,Changsha 410078,China;4.Department of Laboratory Medicine,The Third Xiangya Hospital,Central South University,Changsha 410013,China）

Abstract：OBJECTIVERight ventricular（RV）remodeling is one of the essential pathological features in pulmonary arterial hypertension（PAH）.RV hypertrophy or fibrosis are the leading causes of RV remodeling.Magnolol is a com⁃pound isolated from Magnolia officinalis.It possesses multiple pharmacological activities,such as anti-oxidation and anti-inflammation.This study aims to evaluate the effects and underlying mechanisms of magnolol on RV remodeling in hypoxia-induced PAH.METHODS①Male SD rats（220 g）were randomly divided into 5 groups（n=10）：the normoxia group,the hypoxia group,the hypoxia plus Magnolol（10 and 20 mg·kg-1·d-1）group,and the vehicle group.Rats in the normoxia group were kept in a normoxia environment for 4 weeks,while rats in the hypoxia group were kept in a hypoxic chamber（10%O2）.The rats in the hypoxia plus magnolol groups were administered with magnolol at 10 or 20 mg·kg-1（ip）once a day for 4 weeks.At the end of 4 weeks,the heart function was assessed by Doppler echocardiography,and then the rats were anesthetized with sodium pentobarbital（30 mg·kg-1,ip）.The RVSP was measured by the right heart catheterization method.The heart tissues were collected and dissected to calculate the index of RV remodeling（RV/LV+IVS,RV/tibial length,or RV/body weight）.Part of the RV samples was fixed with 4% paraformaldehyde for morphological analysis,while other samples were frozen at-80℃ for molecular studies（measurements of ANP,BNP,α-SMA,and col⁃lagen Ⅰ/Ⅲ mRNA expression as well as p-JAK2/JAK2 and p-STAT3/STAT3 protein levels）.② To evaluate the effect of magnolol on hypoxia-induced myocardial hypertrophy and fibrosis,H9c2 or cardiac fibroblasts were divided into 7 groups：the control group,cells were cultured under normal conditions;the hypoxia group,cells were cultured under hypoxic condition（3%O2）;the hypoxia plus magnolol 10 mg·kg-1group,magnolol 10μmol·L-1was added to the culture medium before the hypoxia treatment;the hypoxia plus magnolol 30 mg·kg-1group,magnolol 20 μmol·L-1was added to the culture medium before the hypoxia treatment;the hypoxia plus TG-101348 group,TG-101348（a specific inhibitor of JAK2）1 μmol·L-1was added to the culture medium before the hypoxia treatment;the hypoxia plus JSI-124 group,JSI-124（a specific inhibitor of JAK2）1 μmol·L-1was added to the culture medium before the hypoxia treatment;and the hypoxia plus vehicle group,an equal volume of vehicle（DMSO）was added to the culture medium before the hypoxia treatment.At the end of the experiments,the cells were collected for morphological and molecular analysis.RESULTSIn vivo,male Sprang-Daley rats were exposed to 10%O2for 4 weeks to establish an RV remodeling model,which showed hypertrophic and fibrotic features（increases of RV remodeling index,cellular size,hypertrophic and fibrotic marker expression）,accompanied by an elevation in phosphorylation levels of JAK2 and STAT3;these changes were attenuated by treating rats with magnolol.In vitro,the cultured H9c2 cells or cardiac fibroblasts were exposed to 3%O2for 48 h to induce hypertrophy or fibrosis,which showed hypertrophic（increases in cellular size as well as the expression of ANP and BNP）or fibrotic features（increases in the expression of collagen Ⅰ,collagen Ⅲ and α-SMA）.Administration of mag⁃nolol and TG-101348 or JSI-124（JAK2 selective inhibitors）could prevent the process of myocardial hypertrophy and fibrosis,accompanied by the decrease in the phosphorylation level of JAK2 and STAT3.CONCLUSIONMagnolol can attenuate RV hypertrophy and fibrosis in hypoxia-induced PAH rats through a mechanism involving inhibition of the JAK2/STAT3 signaling pathway.

Key words：magnolol;right ventricle remodeling;myocardial hypertrophy;myocardial fibrosis;JAK2;STAT3

Corresponding author：PENG Jun,E-mail：Junpeng@csu.edu.cn;LIU Bin,E-mail：liubin@csu.edu.cn

Corticosterone induced D-serine release deficit play an important role in long-term potentiation impairment by corticosterone in perforant path-dentate gyrus pathway

HUANG Yan1,2,WANG Chen1,2,YU Qi1,2,LI Dong1,2,SUN Na1,2,ZHANG Yong-xiang1,2,ZHOU Wen-xia1,2

（1.Beijing Institute of Pharmacology and Toxicology,Beijing 100850,China;2.State Key Laboratory of Toxicology and Medical Countermeasures,Beijing 100850,China）

Abstract：OBJECTIVEPrevious studies showed that over activation of NMDA receptors may be a crucial cause of long-term potentiation（LTP）and cognitive impairment induced by stress or corticosterone.However,other studies showed that the function of NMDA receptors is insufficient since the NMDA receptors co-agonist D-serine could improve stress-induced cognitive impairment.The purpose of this study is to clarify whether over activation of NMDA receptors or hypofunction of NMDA receptors is involved in hippocampal impairment of LTP by corticosterone and the underlying mechanisms.METHODSCort was injected subcutaneously 1 h before the high-frequency stimulation（HFS）to induce LTP impairment.NMDA receptor antagonists and agonists were administrated by icv.RESULTSHippocampal LTP and object location recognition memory were impaired in corticosterone-treated mice.Corticosterone increased the gluta⁃mate level in hippocampal tissues,neither NMDA receptors antagonist nor its subtype antagonists alleviated impairment of LTP,while enhancing the function of NMDA receptors by D-serine did alleviate impairment of LTP by corticosterone,suggesting that hypofunction of NMDA receptors might be one of the main reasons for impairment of LTP by corticoste⁃rone.Further results showed that the level of D-serine and its precursor L-serine did not change.D-serine release-related protein Na+-independent alanine-serine-cysteine transporter-1（ASC-1）in the cell membrane was decreased and increas⁃ing D-serine release by the selective activator of ASC-1 antiporter activity alleviated impairment of LTP by corticoste⁃rone.CONCLUSIONTaken together,this study demonstrates that hypofunction of NMDA receptors may be involved in impairment of LTP by corticosterone and reduced D-serine release may be an important reason for its hypofunction,which is an important complement to existing mechanisms of corticosterone-induced LTP and cognitive impairment.

Key words：corticosterone;long-term potentiation;N-methyl-D-aspartate receptors;D-serine;Na+-independent ala⁃nine-serine-cysteine transporter-1

Corresponding author：ZHANG Yong-xiang,E-mail：zhangyx@bmi.ac.cn;ZHOU Wen-xia,E-mail：zhouwx@bmi.ac.cn

Influence and mechanism of Xindakang on cardiac functions of isolated frog heart

KONG Xiu-mei,JIANG Yi,HAN Yi-wei,ZHAO Qin

（Engineering Research Center of Tibetan Medicine Testing Technology of Ministry of Education,Tibetan Medicine Screening Laboratory,School of Medicine,Tibet University for Nationalities,Xianyang 712082,China）

Abstract：OBJECTIVETo evaluate the effects of flavonoids from Xindakang（Hippophae Fructus flavone）on myo⁃cardial systolic and diastolic functions of isolated frog hearts and explore the possible mechanism,and provide experi⁃mental basis for improving the effect and efficacy of Xindakang on cardiac function.METHODSThe isolated frog heart perfusion specimens were prepared by Yagi′s method,and the effects of different concentrations of Xindakang on myo⁃cardial contractility（0.0125,0.025,0.05,0.1 and 0.2 g·L-1）,heart rate and cardiac output of isolated frog heart were stud⁃ied.Acetylcholine,atropine and epinephrine were administered successively to analyze the effects of Xindakang on car⁃diac systolic function of isolated frogs under the action of different drugs,and compared with propranolol.The effect of extracellular calcium ion concentration on the action of Xindakang was studied by using low calcium concentration,high cal⁃cium concentration and normal Ren′s solution.To study the effect and possible mechanism of Xindakang on cardiac systolic function of frog.RESULTSThe concentration of Xindakang in the range of 0.0125-0.1 g·L-1could weaken the contractility of isolated frog heart and increase the concentration of Xindakang.The inhibitory effect of Xindakang on con⁃tractility of isolated frog heart was enhanced,and showed obvious dose-effect relationship.Cardiac output was signifi⁃cantly decreased by Xindakang（P＜0.01）,slow heart rate（P＜0.05）;M receptor blocker atropine could not antagonize the contractile effect of Xindakang,and Xindakang could not completely antagonize the contractile effect of adrenalin.Xindakang could inhibit the isolated frog heart in low calcium concentration,high calcium concentration and normal Ren′s solution,and increased with the increase of extracellular calcium concentration（P＜0.01）.CONCLUSIONXinda⁃kang has inhibitory effect on isolated frog heart,which may be achieved by blocking the calcium channel on myocardial cell membrane and reducing the calcium concentration in myocardial cells.

Key words：Hippophae Fructus;flavone;Xindakang;isolated frog heart;cardiac function

Foundation item：Natural Science Foundation of Tibet Autonomous Region（2015ZR-13-16）

Corresponding author：ZHAO Qin,E-mail：xyzhaoqin@126.com

Preliminary study on ecological environment and biological resources of Qinba areas

LI Ben1,ZHANG Xiao-yao1,FENG Si-qi1,ZHANG Xiao-ying1,2,3

（1.Chinese-German Joint Laboratory for Natural Product Research,College of Biological Science and Engineering,Shaanxi University of Technology,Hanzhong 723000,China;2.Centre of Molecular and Environmental Biology,Department of Biology,University of Minho,Braga 4710-057,Portugal;3.Department of Biomedical Sciences,Ontario Veterinary College,University of Guelph,Guelph,Canada）

Abstract：The Qinba areas,located in the Central China,represent 317 000 km2,cover 6 provinces.The major mountain of Qinba areas,the Qinling,separates China North and South.This region is one of the areas with the most bio⁃diversity and ecological importance in China and globally,and considered as natural gene bank and bio-resource bank.Qinba mountain areas,together with the Alps in Europe and the Rocky Mountains in North America,are known as the"Three sisters of the earth"by the geological and biological circles.Most of the lands in this region are protected as national park,natural reserve,water resource protection area,etc.These areas have well preserved ecosystem and breeding and habitat area for many endangered species of plants and animals,inculding Panda.In this region,there are more than 4000 species of higher plants,about 2000 species of medicinal plants and animals,200 species of wild ornamental plants and 180 species of economic fungi with edible and medicinal value in those areas.Medicinal plant cultivation,eco-agricul⁃ture and ecotourism are the major industries in this region.The State Key Laboratory of Biological Resources and Ecological Environment of Qinba Areas,located in Hanzhong,the central city of Qinba,is focusing on pan-regional biological resources research and rational utilization,ecological protection,and comprehensive and coordinated regional development.

Key words：Qinba mountain areas;ecosystem;biological resources;biodiversity;medicinal plant;regional develop⁃ment

Network pharmacology analysis and experimental validation to explore mechanism of Shenlian extract on myocardial ischemia

LI Jing-jing,ZHU Xiao-xin

（Institute of Chinese Materia Medica,China Academy of Chinese Medical Science,Beijing 100700,China）

Abstract：OBJECTIVETo explore the potential molecular mechanism of Shenlian（SL）on myocardial ischemia（MI）on the basis of network pharmacology.METHODSFirstly,the main active ingredients of SL were screened in the Traditional Chinese Medicine Integrated Database,and the MI-associated targets were collected from the DisGeNET database.Then,we used compound-target and target-pathway networks to uncover the therapeutic mechanisms of SL.On the basis of network pharmacology analysis results,we assessed the effects of SL in MI rat model and oxygen glu⁃cose deprivation model of H9c2 cells and validated the possible molecular mechanisms of SL on myocardial injury in vivo and in vitro.RESULTSThe network pharmacology results showed that 37 potential targets were recognized,including TNF-α,Bcl-2,STAT3,PI3K,and MMP2.The pathways revealed that the possible targets of SL were involved in the reg⁃ulation of inflammation and apoptosis signaling pathway.Then,in vivo experiments indicated that SL significantly reduced the myocardial infarction size of MI rats.Serum CK-MB,cTnT,CK,LDH,and AST levels were significantly decreased by SL（P＜0.05 or P＜0.01）.In vitro,SL significantly increased H9c2 cell viability.The levels of inflammation factors including TNF-α and MMP2 were significantly decreased by SL（P＜0.05 or P＜0.01）.TUNEL and Annexin V/propidium iodide assays indicated that SL could significantly decrease the cell apoptotic rate in vivo and in vitro（P＜0.05 or P＜0.01）.The remarkable upregulation of anti-apoptotic Bcl-2 and downregulation of pro-apoptotic Bax protein level further confirmed this result.Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the PI3K-Akt and JAK2-STAT3 pathways were significantly enriched in SL.Compared with the model group,SL treatment significantly activated the PI3K-Akt and JAK2-STAT3 pathways in vivo and in vitro according to Western blotting analyses.CONCLU⁃SIONSL could protect the myocardium from MI injury.The underlying mechanism may be related to the reduction of inflammation and apoptosis by activating the PI3K/Akt and JAK2/STAT3 pathways.

Key word：network pharmacology;myocardial ischemia;Shenlian extract;inflammation;apoptotic

Corresponding author：ZHU Xiao-xin,E-mail：xxzhu@icmm.ac.cn

Pharmacological effects of Schisandra Chinensis Fructus polysaccharide on intestinal protection

LI Ming-song,ZHANG Dan-shen,JING Yong-shuai

（College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China）

Abstract：Schisandra Chinensis Fructus（SCF）is the fruit of Schisandra chinensis（Turcz.）Baill.,a perennial vine.It was first recorded in Shen Nong′s herbal classic and has a long application history.Studies have shown that SCF has anti-inflammatory,protective liver,antioxidant,antibacterial and other pharmacological effects.Ancient prescriptions are commonly used in the treatment of chronic diarrhea and other intestinal diseases and diabetes.Modern clinical phar⁃macology features of SCF polysaccharide（SCFP）in diabetes,liver diseases,enteritis and other aspects have achieved excellent results.Gut is an important digestive organ of human body,but intestinal diseases are varied,including Crohn′s disease,ulcerative colitis,intestinal flora imbalance,etc..It is a chronic and non-specific inflammatory disease.The disease is persisted for a long time and the incidence rate is expected to rise.Most of the symptoms are recurrent diarrhea,bloody stool and abdominal pain.It is considered by the World Health Organization as a refractory disease.At present,there is little possibility of complete cure,which is closely related to complex environmental factors,eating hab⁃its and heredity.In recent years,clinical studies have found that SCFP has a variety of pharmacological effects on intes⁃tinal protection.① Reduce inflammatory factors：intestinal mucositis is a common adverse reaction in patients with chemo⁃therapy.The development of mucositis is related to pro-inflammatory factors such as tumor necrosis factor-α （TNF-α）,interleukin-6（IL-6）,IL-1β,Interferon-γ （IFN-γ）.SCFP can significantly reduce IL-6 TNF-α,IL-1β,and IL-8,as well as the accumulation of T cells in the process of resisting apoptosis,reduce the inflammatory reaction and protect the dam⁃age to villi and crypts,improve the symptoms of small intestinal mucositis caused by weight loss and diarrhea.② Pro⁃mote immunoglobulin A secretion：intestinal mucosal immunity is the first line of defense of the body′s immune system.Its main antibody is secretory immunoglobulin A,which can destroy and phagocytize microorganisms,bacteria and viruses.SCFP can improve intestinal immunity by increasing the number and activity of T lymphocytes,promoting the secre⁃tion of secretory immunoglobulin A,and affecting the activity of a variety of cytokines.③Regulation of intestinal flora：the flora in the intestine has the functions of auxiliary nutrient absorption,biological antagonism and immune regulation,and can form a natural barrier for the host's intestine.When the human intestinal flora is disordered,probiotics will be greatly reduced,harmful bacteria will proliferate and destroy the intestinal environment.Under these conditions,the intake of SCFP significantly increased the number of beneficial bacteria such as bifidobacteria and lactobacillus,and sig⁃nificantly decreased the number of conditional pathogens such as enterococcus and escherichia coli,indicating that SCFP can indeed regulate the intestinal disorder caused by lincomycin hydrochloride to a certain extent.This may be because beneficial bacteria in the intestine metabolize polysaccharides produce short chain fatty acids such as lactic acid and acetic acid,which reduces the pH value in the intestine and inhibits the growth of enterococcus and Escherichia coli.In conclusion,SCFP can treat and protect intestinal diseases to a certain extent,which provides a favorable basis for the treatment of intestinal diseases.

Key words：Schisandra Chinensis Fructus;polysaccharide;intestinal diseases;inflammatory factor

Foundation item：Fourth Chinese Materia Medica Resources Survey（Z135080000022）

Corresponding author：JING Yong-shuai,E-mail：cjys1985@126.com

Pulsatilla chinensis：phytochemistry,pharmacology and new drug development

LIU Mao-lun1,SUN Qiang1,ZENG Sha1,CHEN Li1,ZHAO Hui1,YANG Han1,REN Shan1,MING Tian-qi1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：Pulsatilla chinensis is a widely used traditional Chinese herb,which contains 56 types of chemical constit⁃uents,mainly including triterpenoid saponins,organic acids,coumarins and lignans.The largest portion of the ingredi⁃ents in Pulsatilla chinensis is the family of triterpenoid saponins,in which anemoside B4 is the major effective compound and indexing component.The main components of Pulsatilla chinensis can metabolize into a vast array of active prod⁃ucts in vivo,which play vital roles in its biological activity.Mounting evidence reveals that Pulsatilla chinensis exerts a wide range of therapeutic activities,such as anti-cancer,immunoregulation,anti-inflammation and anti-schistosome,with fewer adverse reactions,via various signaling pathways and multiple targets.It was documented that the active ingre⁃dient of Pulsatilla chinensis can lessen the drug resistance and synergize the effects of other natural products includ⁃ing paclitaxel,as well as ameliorate the clinical efficacy of chemical drugs,such as adriamycin.However,Pulsatilla chi⁃nensis was also reported to be possibly the main cause of hemolysis and chronic liver injury.The efforts should be made to deeply investigate the pharmacological actions and underlying mechanisms of Pulsatilla chinensis,with a focus on the anti-cancer efficacy,and develop new drugs based on the components of Pulsatilla chinensis for future utilization in the clinical setting.

Key words：Pulsatilla chinensis;phytochemistry;pharmacology

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;Excellent Talent Program of Chengdu University of Tra⁃ditional Chinese Medicine of China（YXRC2019002;ZRYY1917）;and Open Research Fund of the State Key Laboratory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Z-Guggulsterone alleviated renal fibrosis and G2/M cycle arrest through Klotho/P53 signaling

LIU Min-na1,LIU Tian-long2

（1.Department of Nephrology,2.Department of Clinical Pharmacy,the 940th Hospital of PLA Joint Logistics Support Forces,Lanzhou 730050,China）

Abstract：OBJECTIVEChronic kidney disease（CKD）has become a global public health problem with 10%-15%incidence rate,and inhibiting the renal interstitial fibrosis is considered to be a potential strategy to delay the progression of CKD.Z-Guggulsterone（Z-GS）,an active compound from derived from Commiphora mukul,has been proved to be effective in various diseases.The present study aimes to determine the protective effect and the molecular mechanism of Z-GS on renal fibrosis.METHODSUnilateral ureteral obstruction（UUO）mice and hypoxia-induced HK-2 cells were used to simulate renal fibrosis in vitro and in vivo,respectively.The mice and cells were treated with different doses of Z-GS to observe the pharmacological action.Renal function,including Scr,BUN,and UA,were detected by commercial kits.H&E and Masson staining were performed to observe histopathological changes of kidney.Cell viability and LDH release of HK-2 cells were detected by commercial kits.Cell cycle distribution and apoptosis rate were analyzed by flow cytometry.Fibrosis markers were detected by immunohistochemistry and immunofluorescence analysis.Cell cycle related proteins and Klotho/p53 signaling were analyzed by Western blotting.RESULTSThe results showed that Z-GS decreased the rise of Scr,BUN,and UA and lightened renal histopathological injury,which were induced by UUO.Besides,Z-GS administration alleviated renal fibrosis in mice by inhibiting the expressions of α-SMA,TGF-β and colla⁃gen Ⅳ,and delayed G2/M cell cycle arrest by promoting the expressions of CDK1 and cyclinD1/B1 rate.Experiments in vitro indicated that Z-GS treatment significantly increased the cell viability while decreased the LDH release in hypoxiainduced HK-2 cells.In addition,hypoxia induced fibrosis and G2/M cycle arrest in HK-2 cells were retarded by Z-GS.The study of its possible mechanism exhibited that Z-GS treatment increased the level of Klotho and inhibited P53 level.Nev⁃ertheless,the effect of Z-GS on Klotho/P53 signaling was reversed by siRNA-Klotho.Moreover,siRNA-Klotho treatment eliminated the effects of Z-GS on G2/M cell cycle arrest and fibrosis.CONCLUSIONThis study clarified that Z-GS allevi⁃ated renal fibrosis and G2/M cycle arrest through Klotho/P53 signaling pathway.People who have suffered CKD may potentially benefit from treatment with Z-GS.

Key words：chronic kidney disease;renal interstitial fibrosis;Z-Guggulsterone;G2/M cycle arrest;Klotho/P53 signaling

Foundation item：National Natural Science Foundation of China（82003982）;Natural Science Foundation of Gansu Province（20JR5RA591;20JR10R015）;and Special Cultivation Project of the 940th Hospital（2021yxky026）

Effects of total flavonoids of Rhododendra simsii on ameliorating brain injury via G protein-coupled SOCE pathway mediated by STIM and Orai in subacute phase of ischemia/reperfusion

LU Jia-jun1,JIANG Chen-chen1,HE Yu-xiang1,SHI Lei1,YIN Xiu-yun1,CHEN Zhuo1,CAO Di1,2,HAN Jun1,2,3

（1.Department of Pharmacology,Drug Research and Development Center,School of pharmacy,Wannan Medical College,Wuhu 241002,China;2.Anhui Provincial Engineering Laboratory for Screening and Re-evaluation of Active Compounds of Herbal Medicines in Southern Anhui,Wuhu 241002,China;3.Anhui Provincial Engineering Research Center for Polysaccharide Drugs,Wuhu 241002,China）

Abstract：OBJECTIVETo explore the effect of total flavonoids of Rhododendra simsii（TFR）on improving cerebral ischemia/reperfusion injury（CIRI）and its relationship with STIM/Orai-regulated operational Ca2+influx（SOCE）pathway.METHODSOxygen-glucose deprivation/reoxygenation（OGD/R）PC12 cells were used to simulate CIRI in vitro,and the intracellular Ca2+concentration and apoptosis rate of PC12 cells were detected by laser confocal microscope and flow cytometry,respectively.The regulation of STIM/Orai on SOCE was analyzed by STIM/Orai gene silencing and STIM/Orai gene overexpression.The CIRI model was established by MCAO in SD rats.The activities of inflammatory cyto⁃kines IL-1,IL-6 and TNF-α in serum were detected by ELISA.The pathological changes of ischemic brain tissue and the infarction of rat brain tissue were detected by HE staining and TTC staining.The protein and mRNA expression levels of STIM1,STIM2,Orai1,caspase-3 and PKB in brain tissue were detected by Western blotting and RT-qPCR,respectively.RESULTSThe results of in vitro experiment showed that the fluorescence intensity of Ca2+and apoptosis rate in PC12 cells treated with TFR were significantly lower than those in OGD/R group,and this trend was enhanced by SOCE antagonist 2-APB.STIM1/STIM2/Orai1 gene silencing significantly reduced apoptosis and Ca2+overload in OGD/R model,while TFR combined with overexpression of STIM1/STIM2/Orai1 aggravated apoptosis and Ca2+overload.In the in vivo experiment,TFR significantly reduced the brain histopathological damage,infarction of brain tissue,the contents of IL-1,IL-6 and TNF-α in the serum in MCAO rats and down-regulated the expression of STIM1,STIM2,Orai1 and caspase-3 protein and mRNA in the brain tissue,and up-regulated the expression of PKB.The above effects were enhanced by the addition of 2-APB.CONCLUSIONThe above results indicate that TFR may reduce the contents of inflammatory factors and apoptosis,decrease Ca2+overload and ameliorate brain injury by inhibiting SOCE pathway mediated by STIM and Orai,suggesting that it has a protective effect against subacute CIRI.

Key words：total flavonoids of Rhododendra simsii;cerebral ischemia/reperfusion injury;STIM/Orai;store-operated calcium entry;2-APB

Foundation item：National Natural Science Foundation of China（81173596）;and Major Project of Natural Science Foundation of the Department of Education of Anhui Province（KJ2019ZD32）

Corresponding author：HAN Jun,E-mail：aku110@153.com

Study on potential mechanism of hyperoside on improving ischemia/reperfusion injury based on network pharmacology

LU Jia-jun1,JIANG Chen-chen1,SHI Lei1,CAO Di1,3,HAN Jun1,2,3

（1.Department of Pharmacology,Drug Research and Development Center,School of pharmacy,Wannan Medical College,Wuhu 241002,China;2.Anhui Provincial Engineering Laboratory for Screening and Re-evaluation of Active Compounds of Herbal Medicines in Southern Anhui,Wuhu 241002,China;3.Anhui Provincial Engineering Research Center for Polysaccharide Drugs,Wuhu 241002,China）

Abstract：OBJECTIVETo predict the potential targets of hyperoside（Hyp）on improving ischemia/reperfusion injury by network pharmacology,and explore its possible mechanism combined with related literature.METHODSThe action targets of Hyp and ischemia/reperfusion injury were obtained by TCMSP,Swiss Target Prediction,Pharm Mapper,Simi⁃larity ensemble approach,Online Mendelian Inheritance in Man,DisGENT and database.The common targets of drugs and diseases were screened by Omishare and STRING database respectively,and the protein-protein interaction（PPI）network map was constructed.Then the interaction network between Hyp and disease targets was constructed by Cyto⁃scape software and topological cross-linking analysis was carried out.Then the interaction network between Hyp and disease targets was constructed and cross-linked analysis was carried out by using Cytoscape software.The gene ontol⁃ogy（GO）of the core target was analyzed by David database,and then the related pathways of the core target were enriched by KEGG database.RESULTSA total of 54 GO enrichment processes were obtained by GO enrichment anal⁃ysis of 44 common genes,including 38 biological processes（BP）,15 cell composition（CC）processes,and 1 molecular functional（MF）process.43 items were obtained by signal pathway enrichment analysis in KEGG database.CONCLU⁃SIONIt is suggested that the mechanism of Hyp may be related to PI3K-Akt,RAP1,RAS,VEGF and other signal trans⁃duction pathways.The above results laid a theoretical foundation for the study of the mechanism and clinical application of the treatment of ischemia/reperfusion injury.

Key words：hyperoside;ischemia/reperfusion injury;network pharmacology

Foundation item：National Natural Science Foundation of China（81170148）;and Major Project of Natural Science Foundation of the Department of Education of Anhui Province（KJ2019ZD32）

Corresponding author：HAN Jun,E-mail：aku110@163.com

Progress in protective effect and mechanism of 6-gingerol on myocardial ischemia/reperfusion injury

MA Yun-feng1,PAN Fei-bing2,ZHANG Dan-shen1,JING Yong-shuai1

（1.College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China;2.Huachuang Institute of Areca Research-Hainan,Haikou 570125,China）

Abstract：The morbidity and mortality of cardiovascular diseases are very high,which has attracted more and more attention all over the world.Common treatment methods for clinical treatment of acute myocardial infarction include direct percutaneous coronary intervention and coronary artery bypass grafting,which can quickly restore blocked coronary blood flow and reduce the infarct size.However,the inevitable ischemia/reperfusion injury will occur during the recovery of coronary blood flow,its pathological mechanism is complicated,and the Western medicine countermeasures are very limited.Among the current drugs for the treatment of cardiovascular diseases,traditional Chinese medicine has become a research hotspot due to its multiple targets,safety,and low side effects.Ginger is the fresh rhizome of Zingiber offici⁃nale Rosc.,a perennial herbaceous plant in the ginger family.It is a dual-purpose resource of medicine and food.Ginger has the functions of relieving the appearance and dispelling cold,warming up and relieving vomiting,resolving phlegm and relieving cough,and relieving fish and crab poison.The chemical components of ginger mainly include volatile oil,gingerol,diphenylheptane,etc..Among them,6-gingerol,as the main active component of gingerols,has obvious phar⁃macological effects in myocardial protection,anti-oxidation,anti-inflammatory,etc..Studies have shown that 6-gingerol protects myocardium mainly through anti-oxidative stress,anti-inflammatory,inhibiting cell apoptosis,and preventing cal⁃cium influx.①Anti-oxidative stress：oxidative stress is a state where oxidation and anti-oxidation in the body are out of balance,and it is also an important factor leading to myocardial damage.Many studies have confirmed that 6-gingerol has an antioxidant effect,and it is considered a natural antioxidant.6-gingerol can significantly reduce the degree of oxi⁃dative stress and the level of reactive oxygen species caused by cardiomyocyte damage,and has a significant cardiopro⁃tective effect.②Anti-inflammatory：inflammation can cause substantial cell damage and organ dysfunction,which is another important cause of myocardial damage.6-gingerol can reduce the levels of inflammatory factors such as inter⁃leukin-6,interleukin-1β,and tumor necrosis factor-α in cardiomyocytes,and at the same time inhibit the TLR4/NF-κB sig⁃naling pathway,an important regulatory pathway of inflammation,showing that it may improve myocardial damage through anti-inflammatory effects.③ Inhibition of apoptosis：apoptosis is a complex and orderly process in the autono⁃mous biochemical process of cells,and one of the main mechanisms of myocardial injury.This process can be roughly divided into three pathways：mitochondria,endoplasmic reticulum,and death receptors.Among them,the mitochondrial pathway plays an important role,and Bcl-2 and Bax located upstream of this pathway can regulate the entire process of cell apoptosis by regulating the permeability of the mitochondrial membrane.Studies have found that the preventive application of 6-gingerol can reduce cell damage,reduce the number of apoptotic cells,reduce the activity of Bax and caspase-3,and increase the expression of Bcl-2.Therefore,6-gingerol pretreatment can reduce the damage of cardio⁃myocytes,and its mechanism may be related to the inhibition of apoptosis.④Prevent calcium influx：calcium overload is involved in the pathogenesis of myocardial ischemic injury,which may be related to excessive contracture,arrhythmia,and mitochondrial Ca2+accumulation that impairs myocardial function.6-gingerol inhibits the increase of intracellular Ca2+concentration by inhibiting L-type calcium current,thereby reducing extracellular Ca2+influx,thereby avoiding calcium overload and playing a cardioprotective effect.In summary,6-gingerol can effectively treat and improve myocardial isch⁃emia/reperfusion injury,and it has great development potential in the fields of medicine and health products.

Key words：6-gingerol;myocardial ischemia/reperfusion injury

Foundation item：Fund of Dean of Huachuang Institute of Areca Research-Hainan（HCBL2020YZ-012）

Corresponding author：JING Yong-shuai,E-mail：cjys1985@126.com

Research advances in phytochemistry, pharmacology and toxicology of oleanolic acid

REN Shan1,SUN Qiang1,CHEN Li1,ZENG Sha1,ZHAO Hui1,LIU Mao-lun1,YANG Han1,MING Tian-qi1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：Oleanolic acid（OA）is a pentacyclic triterpenoid chemical component that exists in natural plants with a molecular formula of C30H48O3and a molecular weight at 456.71 g·mol-1.OA is widespread in traditional Chinese herbal medicine（Ligustri Lucidi Fructus,Achyranthis Bidentate Radix,Red Sage）and berries（blueberries,grapes）.In recent years,because of the extensive pharmacological effects of OA,its advantages in disease treatment have become increasingly prominent and gradually attracted the attention of pharmaceutical researchers.OA has effective therapeutic effects on a series of chronic diseases such as inflammation,cancer,diabetes,and cardiovascular diseases through mul⁃tiple signaling pathways and various targets.Especially in cancers,such as colorectal cancer,liver cancer,gastric cancer,lung cancer,breast cancer and other malignancies,OA presents substantial efficacy.However,its poor aqueous solubility,needy bioavailability,and unsatisfactory pharmacological activity excessively restrict its clinical application.More impor⁃tantly,the improper utilization of OA can cause adverse reactions,toxic effects and even damage to organs in some spe⁃cific situations.With the discovery of various pharmacological effects,the complex action mechanisms of OA,the contin⁃uous progress in structural modification of OA,as well as the synthesis of OA derivatives,its application is expand⁃ing gradually.Among numerous studies,there is a clear indication that OA and its derivatives,if fully developed,may provide an alternative and cheaper treatment for a variety of chronic diseases.However,the specific molecular mecha⁃nisms of OA and its derivatives as an alternative therapy and supplementary therapy for cancer,diabetes,cardiovascular disease and other chronic diseases remain to be clarified.Therefore,it is necessary to further study the pharmacokinet⁃ics,pharmacological activity,specific targets and related mechanisms of OA to lay a solid foundation for drug devel⁃opment and the application of OA in clinical settings.

Key words：oleanolic acid;pharmacology;toxicology;derivatives;review

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;Excellent Talent Program of Chengdu University of Tra⁃ditional Chinese Medicine of China（YXRC2019002;ZRYY1917）;and Open Research Fund of the State Key Laboratory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Inhibitory action of berberine on colorectal cancer HCT116 cells by regulation of Hedgehog signaling pathway

SUN Qiang1,CHEN Li1,ZENG Sha1,LIU Mao-lun1,REN Shan1,ZHAO Hui1,YANG Han1,MING Tian-qi1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVETo investigate the inhibition and mechanism of berberine on human colorectal cancer HCT116 cells through canonical Hedgehog signaling pathway.METHODSThe effect of berberine on cell morphology was observed by microscopy.MTT colorimetric assay,cell scratch experiment,colony formation assay and Hoechest/PI staining were utilized to detect the activities of berberine on cell viability,cell migration and cell apoptosis.Flow cytome⁃try was applied to examine the cell apoptosis.The effects of berberine on caspase-3 and caspase-9 were detected by caspase activity detection kit.The expressions of Hedgehog signaling pathway-related proteins SHH,GLI1,PTCH1,SMO,SUFU,apoptosis-related proteins Bax and Bcl-2 as well as cell cycle-related proteins cyclin D1 were detected by Western blotting.Additionally,quantitative real time RT-PCR was employed to assess the mRNA expression levels of Hedgehog signaling pathway-related genes SHH,GLI1,PTCH1,SMO,SUFU,apoptosis-related genes Bax and Bcl-2 as well as cell cycle-related genes cyclin D1.RESULTSBerberine sharply altered the morphology of human colorectal cancer HCT116 cells,demonstrated by that migration ability of HCT116 cells was reduced significantly and the nuclei were densely stained.Berberine could induce apoptosis in a dose-dependent manner.The activities of caspase-3 and caspase-9 were increased prominently.The expression levels of Hedgehog signaling pathway-related protein SUFU and apoptosis-related protein Bax were augmented substantially.The expression levels of Hedgehog signaling pathway related proteins SHH,GLI1,PTCH1,SMO,apoptosis-related protein Bcl-2 as well as cell cycle-related genes cyclin D1 were markedly lessened.Besides,the mRNA expression levels of Hedgehog signaling pathway-related gene SUFU and apoptosis-related gene Bax were augmented substantially.The mRNA expression levels of Hedgehog signaling path⁃way-related genes SHH,GLI1,PTCH1,SMO,apoptosis-related gene Bcl-2 as well as cell cycle-related gene cyclin D1 were markedly lessened.CONCLUSIONBerberine,which is the main component of coptidis rhizoma,can remarkably restrain the growth and proliferation,promote apoptosis of human colorectal cancer cells HCT116,and the underlying mechanism may be involved in suppressing the activity of the Hedgehog signaling pathway.

Key words：berberine;Hedgehog signaling pathway;colorectal cancer;cell proliferation;cell apoptosis

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;Excellent Talent Program of Chengdu University of Tra⁃ditional Chinese Medicine of China（YXRC2019002;ZRYY1917）;and Open Research Fund of State Key Laboratory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Pathogenesis of chronic social defeat stress model induced depressive-like mouse model according to LC-MS/MS-based metabolomics

WANG Qi1,3,4,XIANG Huan2,ZHAO Hui-liang1,3,4,LING-HU Ting1,3,4,TIAN Jun-sheng1,3,4,QIN Xue-mei1,3,4

（1.Modern Research Center for Traditional Chinese Medicine,2.Department of Physical Education,3.Key Laboratory of Effective Substances Research and Utilization in TCM of Shanxi Province,4.Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education,Shanxi University,Taiyuan 030006,China）

Abstract：OBJECTIVETo explore the pathogenesis of depression according to the LC-MS/MS-based metabolo⁃mics in the mouse model which exhibits social avoidance state induced by the chronic social defeat stress model（CSDS）.METHODSTwenty male C57BL/6N mice were randomly divided into control group and model group suffering CSDS,and the ICR retired breeder mice were used to attack the model group for 14 d of chronic social defeated stress.The open field test and source preference test were both used to observe depression-like behavior.Besides,the social inter⁃action test is used to observe the social interaction state,especially.After the stress,the serum samples of mice were collected,and the changes of endogenous metabolites were analyzed by LC-MS metabolomics technology,and the pathway analysis of the differential metabolites was performed to explore the pathogenesis of the CSDS induced depres⁃sive-like mouse model.RESULTSAfter the stress of CSDS was completed,the mice in the model group showed a significant slowdown in body weight growth,a reduction in the source preference rate,and a significant reduction in the total distance and the number of rearing in the open field test.Distinctively,the social interaction rate is remarkably decreasing.There are 24 differential metabolites found in the serum of CSDS model mice.CONCLUSIONThe mouse who suffered CSDS stress would show depressive-like behavior.Based on the LC-MS/MS metabolomics,24 differential metabolites were found in the serum of CSDS model mice.The amino acid metabolism might be significant to the patho⁃genesis of the CSDS induced depressive-like mouse model.

Key words：depression;chronic social defeat stress model;social interaction test;LC-MS/MS;metabolomics

Corresponding author：QIN Xue-mei,E-mail：qinxm@sxu.edu.cn;TIAN Jun-sheng,E-mail：jstian@sxu.edu.cn

Organ specific differences in alteration of aquaporin expression in rats treated with sennoside A,senna anthraquinones and rhubarb anthraquinones

WANG Zhao-yang,CHENG Ying,QIAO Bo-ling

（Key Laboratory of Resource Biology and Modern Biotechnology in Western China,Ministry of Education,Northwestern University,Xi′an 710069,China）

Abstract：OBJECTIVESenna and rhubarb are classified as stimulative laxatives,and known to have similar effec⁃tive constituents,the anthraquinones.Being protected by the β-glucoside bond,the anthraquinones can reach the intes⁃tines where they are degraded into complex metabolites by enzymes secreted from the intestinal microbiome.It is these complex metabolites that produce the laxative effects.Then the similarities and differences of action between the anthra⁃quinones require further elucidation.METHODSHere,we studied metabolites of senna anthraquinones（SAQ）,rhubarb anthraquinones（RAQ）and their chemical marker,sennoside A（SA）,in a rat diarrhea model.In the in vitro biotransfor⁃mation experiments,SAQ,RAQ and SA were incubated with rat fecal flora solution and the metabolites produced were analyzed using HPLC.In the in vivo studies,the same compounds were investigated for purgation induction,with mea⁃surement of histopathology and multiple aquaporins（Aqps）gene expression in six organs.RESULTSSAQ and RAQ had similar principal constituents but could be degraded into different metabolites.A similar profile of Aqps down-regula⁃tion for all compounds was seen in the colon,suggesting a similar mechanism of action for purgation.However,in the kidneys and livers of the diarrhea-rats,down-regulation of Aqps was found in the RAQ-rats whereas up-regulation of Aqps was seen in the SAQ-rats.Furthermore,the RAQ-rats showed lower aquaporin 2（Aqp2）protein expression in the kidneys,whilst the SA-rats and SAQ-rats had higher Aqp2 protein expression in the kidneys.This may have implications for side effects of SAQ or RAQ in patients with chronic kidney or liver diseases.CONCLUSIONSAQ and RAQ showed similar laxative actions with a similar mechanism,they could display different actions in rat kidneys and livers.We suggest that the clinical usage of senna or rhubarb products should be clarified for patients having chronic kidney or liver diseases.

Key words：senna anthraquinones;rhubarb anthraquinones;sennoside A;laxative;aquaporins

Corresponding author：QIAO Bo-ling,E-mail：bolingq@nwu.edu.cn

Active fraction combination from Liuwei Dihuang decoction improves adult hippocampal neurogenesis and neurogenic microenvironment in cranially irradiated mice

WEI Ming-xiao1,2,FENG Shu-fang3,SHI Tian-yao2,ZHOU Wen-xia2,ZHANG Yong-xiang1,2

（1.School of Life Science and Biopharmaceutics,Shenyang Pharmaceutical University,Shenyang 110016,China;2.State Key Laboratory of Toxicology and Medical Countermeasures,Beijing Institute of Pharmacology and Toxicology,Beijing 100850,China;3.Department of Poisoning and Treatment,Affiliated Hospital to Academy of Military Medical Sciences,Beijing 100039,China）

Abstract：OBJECTIVECranial radiotherapy is clinically used in the treatment of brain tumors;however,the conse⁃quent cognitive and emotional dysfunctions seriously impair the life quality of patients.LW-AFC,an active fraction combi⁃nation extracted from classical traditional Chinese medicine prescription Liuwei Dihuang decoction,can improve cogni⁃tive and emotional dysfunctions in many animal models;however,the protective effect of LW-AFC on cranial irradiationinduced cognitive and emotional dysfunctions has not been reported.Recent studies indicate that impairment of adult hippocampal neurogenesis（AHN）and alterations of the neurogenic microenvironment in the hippocampus constitute crit⁃ical factors in cognitive and emotional dysfunctions following cranial irradiation.Here,our research further investigated the potential protective effects and mechanisms of LW-AFC on cranial irradiation-induced cognitive and emotional dys⁃functions in mice.METHODSLW-AFC（1.6 g·kg-1）was intragastrically administered to mice for 14 d before cranial irra⁃diation（7 Gy γ-ray）.AHN was examined by quantifying the number of proliferative neural stem cells and immature neu⁃rons in the dorsal and ventral hippocampus.The contextual fear conditioning test,open field test,and tail suspension test were used to assess cognitive and emotional functions in mice.To detect the change of the neurogenic microenvi⁃ronment,colorimetry and multiplex bead analysis were performed to measure the level of oxidative stress,neurotrophic and growth factors,and inflammation in the hippocampus.RESULTSLW-AFC exerted beneficial effects on the contex⁃tual fear memory,anxiety behavior,and depression behavior in irradiated mice.Moreover,LW-AFC increased the num⁃ber of proliferative neural stem cells and immature neurons in the dorsal hippocampus,displaying a regional specificity of neurogenic response.For the neurogenic microenvironment,LW-AFC significantly increased the contents of superox⁃ide dismutase,glutathione peroxidase,glutathione,and catalase and decreased the content of malondialdehyde in the hippocampus of irradiated mice,accompanied by the increase in brain-derived neurotrophic factor,insulin-like growth factor-1,and interleukin-4 content.Together,LW-AFC improved cognitive and emotional dysfunctions,promoted AHN preferentially in the dorsal hippocampus,and ameliorated disturbance in the neurogenic microenvironment in irradiated mice.CONCLUSIONLW-AFC ameliorates cranial irradiation-induced cognitive and emotional dysfunctions,and the underlying mechanisms are mediated by promoting AHN in the dorsal hippocampus and improving the neurogenic micro⁃environment.LW-AFC might be a promising therapeutic agent to treat cognitive and emotional dysfunctions in patients receiving cranial radiotherapy.

Key words：Liuwei Dihuang decoction;adult hippocampal neurogenesis;LW-AFC;cognitive and emotional dysfunc⁃tions;cranial irradiation

Corresponding author：ZHANG Yong-xiang,E-mail：zhangyx@bmi.ac.cn;ZHOU Wen-xia,E-mail：zhouwx@bmi.ac.cn;SHI Tian-yao,E-mail：tianyao_shi@126.com

Integrating network pharmacology and pharmacological evaluation for deciphering the mechanism of（-）-epigallocatechin-3-gallate alleviating ethanol-induced endothelial cells injury

XU Jie,ZHANG Shi-hao,YAN Yu,HAO Ting,XU Shou-zhu

（Department of Public Health,Shaanxi University of Chinese Medicine,Xianyang 712046,China）

Abstract：ObjectiveTo investigate the potential therapeutic targets and pharmacological mechanism of（-）-epigal⁃locatechin-3-gallate（EGCG）based on network pharmacology and experimental verification.METHODSThe druggability of EGCG was measured by the traditional Chinese medicine systems pharmacology（TCMSP）server,and potential tar⁃gets of EGCG were identified by Pharm Mapper and Drug Repositioning and Adverse drug Reaction via Chemical-Pro⁃tein Interactome（DRAR-CPI）.The potential targets were imported into GeneMANIA database to obtain the protein-pro⁃tein direct interaction network,and target physical interaction,co-expression,prediction,genetic interaction,and shared protein domains.The biological process,molecular functions,cellular components and KEGG signaling pathways of potential targets were analyzed using DAVID database.For further study,ethanol was used to establish a model of endothelial injury in vitro.The cell viability was assayed by MTT method,the cellular apoptosis was stained by Annexin V/PI,and the expression levels of Bcl-2,Bax and cleved-caspase-3 were tested by Western blotting.Then,JC-1 and nuclear translocation of NF-κB experiments were used to study the mitochondrial membrane potential and nuclear trans⁃location.RESULTSThe oral availability of EGCG was 55.09%（≥ 30%）and drug-like index was 0.77（≥ 0.18）,which were considered pharmacokinetically active.17 potential targetable proteins of EGCG were predicted by Pharm Mapper and DRAR-CPI.Further research showed that 68.13%displayed similar co-expression characteristics,26.11%physical interactions,and 2.74%shared the same protein domain.The depth network analysis results showed that the biofunc⁃tions of EGCG were mainly by regulating glutathione derivative biosynthetic process,glutathione metabolic process,nitrogen compound metabolic process etc..via drug binding,catalytic activity,glutathione transferase activity,anion bind⁃ing etc..in sarcoplasmic reticulum,spindle pole,microtubule cytoskeleton and cytoplasm.KEGG enrichment analysis showed that Glutathione metabolism,IL-17 signaling pathway,EGFR tyrosine kinase inhibitor resistance,PI3K-Akt sig⁃naling pathway and other pathways were involves in the biofunction of EGCG.The above analyses indicated that EGCG exerts its biofunction through antioxidant and anti-inflammatory mechanisms.The experimental results showed that etha⁃nol 20.0 mmol·L-1decreased cell viability,Bcl-2 expression,and increased cell apoptosis,the intracellular ROS,as well as the expression of Bax and cleaved-caspase-3 of human endothelial cells.However,treatment of the cells with EGCG can significantly alleviate ethanol induced endothelial cells injury.Further study showed that EGCG significantly allevi⁃ates ethanol induced mitochondrial depolarization and nuclear translocation of NF-κB.CONCLUSIONS EGCG exerts pharmacological efficacies on ethanol induced endothelial cell injury through multi-target,multi-function and multi-path⁃way mode.Protective effect of EGCG on ethanol induced cell injury was mainly through alteration of mitochondrial func⁃tion and NF-κB translocation.Therefore,EGCG have great potential in protecting against endothelial dysfunction of the persons who are chronically abuse of ethanol.This study also provides a new understanding of EGCG in clinical applica⁃tion on cardiovascular and cerebrovascular diseases.

Key words：（-）-epigallocatechin-3-gallate;ethanol;network pharmacology;target fishing;nuclear translocation of NF-κB

Foundation item：National Natural Science Foundation of China（82100488）;Key Research and Development Pro⁃gram Project of Shaanxi Province（2021SF-071）;and National Training Program of Innovation and Entrepreneurship for Students of China（201910716019;201910716020;202110716027）

Corresponding author：XU Shou-zhu,E-mail：xsz@sntcm.edu.cn

Study on pharmacological effect of Chuanxiong Rhizoma on vascular neuropathic headache

YAN Meng,ZHANG Dan-shen,JING Yong-shuai

（College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China）

Abstract：Chuanxiong Rhizoma is the dry rhizome of Ligusticum chuanxiong in the umbelliferae family.Chuanxiong Rhizoma pungent,warm,go to liver,gallbladder and pericardium.Effective in promoting blood circulation,promoting Qi,dispelling wind and relieving pain,it could treat chest pain,tingling pain in chest and flank,lump,irregular menstruation,amenorrhea,symptomatic abdominal pain,headache and rheumatic pain.Neurovascular headache is a primary disease caused by dysregulation of intracranial vascular movement and nerve function.It has the characteristics of long course,intermittent recurrent attacks,lingering and difficult to heal.Attacks are often accompanied by many plant nervous sys⁃tem symptoms,such as rapid breathing,accelerated heart rate,vomiting,and gastrointestinal dysfunction.Vascular nerve headache is a common clinical disease,frequently bidity.Studies have shown that Chuanxiong Rhizoma has good pharmacological effects in the treatment of vascular neuropathic headache.① The action of Qi and blood circula⁃tion：vascular and neurovascular headache is caused by the evil of external wind and cold and damp heat,which leads to the disconnection of the veins,the disorder of Qi and blood,the obstruction of Qi and blood channels,the loss of brain collateral,and finally causes migraine.Modern Chinese medicine points out that"wind,blood stasis,deficiency,phlegm"are the key factors of the disease.Chuanxiong Rhizoma is the medicine of Qi in the blood.It is pungent and warm.It is good at activating blood and promoting Qi,dispelling wind,relieving pain and dispelling cold,so as to achieve the effect of treating vascular headaches.②Improve brain circulation：angioneurotic headache is caused by dysfunction of the central nervous system related to the regulation of vascular movement,which causes vasospasm or extreme vasodi⁃lation,and the decrease of intracranial blood flow causes cerebral ischemia and hypoxia.Sodium ferulate is a chemical component in Chuanxiong Rhizoma.It has a relatively good inhibitory effect on platelet aggregation and the release of 5-HT from platelets.It can ensure the normal contraction of intracranial and extracranial blood vessels,improve the patient′s brain circulation and nerve function,so as to achieve the effect of treating angioneurotic headaches.③Sedative and analgesic effect：the volatile oil and water decoction of Chuanxiong Rhizoma have sedative and analgesic effects,and the water decoction can counteract the excitatory effect of caffeine.Studies have shown that the ATP activa⁃tion current of rat dorsal root ganglion neurons can be inhibited by ligustrazine in a non-competitive way,which also indi⁃cates that Chuanxiong Rhizoma has a good analgesic effect.In this study,the effects of Chuanxiong Rhizoma on angoneeurotic headache were reviewed,and the pharmacological effects of Chuanxiong Rhizoma were further elucidated,providing basis for clinical application and new drug development of Chuanxiong Rhizoma in the treatment of angoneeu⁃rotic headache.

Key words：Chuanxiong Rhizoma;neurovascular headache;mechanism of action

Foundation item：The S&T Program of Hebei Province（H2021208007）

Corresponding author：JING Yong-shuai,E-mail：cjys1985@126.com

Scutellarin induces apoptosis of colorectal cancer HCT116 cells via Hippo signaling pathway

YANG Han1,SUN Qiang1,ZENG Sha1,CHEN Li1,ZHAO Hui1,LIU Mao-lun1,REN Shan1,MING Tian-qi1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVETo investigate the effect of scutellarin on the apoptosis of human colorectal cancer cells via the Hippo signaling pathway in vitro.METHODSMTT colorimetric method was used to detect the influence of scutellar⁃in on the survival rate of HCT116 cells.And the effect of scutellarin at various concentrations on cell morphology was observed by microscopy.Cell scratch experiment was used to detect the influence of scutellarin on the migration of HCT116 cells.Hoechst 33342/PI double staining method was used to detect the effect of scutellarin on the apoptosis of HCT116 cells.Western blotting method was used to assess the action of scutellarin on the expressions of Hippo signal⁃ing pathway-related proteins Mst1,Lats1,YAP1,p-YAP（Ser127）,TAZ,and its downstream effector proteins c-Myc and cyclin D1,as well as apoptosis-related proteins Bcl-2 and Bax in HCT116 cells.RESULTSScutellarin significantly affected the morphology of HCT116 cells and reduced the survival rate of HCT116 cells.Hoechst 33342/PI double stain⁃ing showed that scutellarin effectively induced the apoptosis of HCT116 cells.Western blotting analysis showed that the expression levels of Hippo signaling pathway-related proteins Mst1,Lats1,YAP1,TAZ and its downstream effector pro⁃teins c-Myc,cyclin D1 were down-regulated in a concentration-dependent manner by scutellarin,and the expression of p-YAP（ser127）was up-regulated.Moreover,scutellarin substantially lessened the expression level of apoptosis-related protein Bcl-2,and promoted the protein level of Bax.CONCLUSIONScutellarin may inhibit the proliferation and migra⁃tion of HCT116 cells,while induce its apoptosis,potentially by activation of Hippo signaling pathway.

Key words：scutellarin;colorectal cancer;HCT116 cells;Hippo signaling pathway

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;Excellent Talent Program of Chengdu University of Tra⁃ditional Chinese Medicine of China（YXRC2019002;ZRYY1917）;and Open Research Fund of the State Key Laboratory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Research progress in mechanism of curcumin in treatment of Parkinson disease

YUAN Xin-ru,JING Yong-shuai,ZHANG Dan-shen

（College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China）

Abstract：Curcumin（Cur）is an important bioactive component of polyphenols in the rhizomes of Curcuma longa L.,Tulipa gesneriana L.and other Curcuma plants.It has a wide range of pharmacological effects such as anti-tumor,antiatherosclerosis,anti-inflammatory,and neuroprotection.Parkinson disease（PD）is a neurodegenerative disease that often occurs in the elderly.Its main pathological characteristics are the characteristic loss of substantia nigra dopaminer⁃gic neurons,the decrease of dopamine content in the striatum,and the formation of Lewy bodies.At present,the main methods of clinical treatment of PD include drug therapy and surgical operation,but due to its complicated pathogene⁃sis,they can only play a role in relieving,but cannot be completely cured.Modern pharmacological studies have shown that Cur has certain effects in the treatment of PD.①Anti-oxidative stress：oxidative stress is closely related to the degeneration of dopaminergic neurons.Studies have found that Cur can increase the activity of superoxide dismutase（SOD）and glutathione peroxidase（GSH-Px）,reduce malondialdehyde（MDA）content,thereby reducing oxidative stress damage and protecting dopaminergic neuron.② Reduce inflammation in brain tissue：neuroinflammation plays an impor⁃tant role in the development of PD.Reducing the level of inflammatory factors can have a certain therapeutic effect on PD.Studies have shown that high-dose Cur can reduce the levels of interleukin-6（IL-6）,IL-1β,and tumor necrosis fac⁃tor-α （TNF-α）in brain tissue,reduce inflammation,inhibit further neuronal damage,improve learning and memory,and exert neuroprotective effects.③ Activation of autophagy：the abnormal accumulation of α-Synuclein（α-Syn）in Lewy bodies is closely related to PD,and autophagy dysfunction leads to α-Syn clearance obstacles and an important factor of abnormal aggregation.Cur can increase the expression of microtubule-associated protein 1 light chain 3（LC3-Ⅱ）and lysosome-associated membrane protein 2A（LAMP2A）,and reduce the protein and mRNA expression of α-Syn.It can be seen that Cur promotes the elimination of α-Syn and protects neurons from damage by activating autophagy.④ Inhi⁃bition of mitochondrial dysfunction：mitochondria plays a central regulatory role in the process of cell apoptosis,and mito⁃chondrial dysfunction is related to reactive oxygen species,energy and mitochondrial membrane potential,which may cause substantia nigra striatal neuropathy.Experiments have shown that Cur can reduce the active oxygen content in PC12 cells induced by MPP+,maintain the normal membrane potential of mitochondria,thereby stabilizing mitochondrial function and inhibiting PC12 cell apoptosis.This study summarized the action mechanism of Cur in the treatment of PD,and clarified the basis of its pharmacodynamics,providing a reference for the clinical research and new drug develop⁃ment research of Cur in the treatment of PD.

Key words：curcumin;Parkinson disease;mechanism of action

Foundation item：Key Research and Development Program of Hebei Province（20370509D）;and Science and Technology Project of Hebei Education Department（ZD2020117）

Corresponding author：ZHANG Dan-shen,E-mail：zhangds2011@126.com

Paeoniflorin-6′-O-benzene sulfonate ameliorates progression of adjuvant-induced arthritis by inhibiting interaction between Ahr and GRK2 of fibroblast-like synoviocytes

ZHANG Bin-jie,WANG Yue-ye,JIA Cheng-yan,LI Su-su,WANG Xin-wei,XU Yuan,CHEN A-yuan,XU He-peng,WANG Chun,WEI Wei,CHANG Yan

（Key Laboratory of Anti-Inflammatory and Immune Medicine,Anhui Collaborative Innovation Center of Anti-inflammatory and Immune Medicine,Institute of Clinical Pharmacology,Anhui Medical University,Ministry of Education,Hefei 230032,China）

Abstract：OBJECTIVEAryl hydrocarbon receptor（Ahr）is thought to be a crucial factor that regulates immune responses,which may be involved in the pathogenesis of autoimmune inflammation including rheumatoid arthritis（RA）.The results of our group in recent years have shown that CP-25,a novel ester derivative of paeoniflorin,has a good effect on improving RA animal models.However,whether the anti-arthritis effect of CP-25 is related to Ahr remains unclear.METHODSCP-25 treatment ameliorated adjuvant-induced arthritis（AA）,a mouse model of RA,by inhibiting Ahr-related activities in fibroblasts like synoviocytes（FLS）.AA rats were treated with CP-25 or paroxetine from day 17 to 33 after immunization.RESULTSCP-25 alleviated arthritis symptoms and the pathological changes,decreased the expression of Ahr in the synovium and FLS of AA rats.Besides,treatment with CP-25 reduced the proliferation and migration of MH7A caused by Ahr activation.In addition,we also demonstrated that CP-25 down-regulated the co-expres⁃sion and co-localization of Ahr and G protein-coupled receptor kinase 2（GRK2）in MH7A.CONCLUSIONThe data pre⁃sented here demonstrated that CP-25 suppressed FLS dysfunction in rats with AA,which were associated with reduced Ahr activation and the interaction between Ahr and GRK2.

Key words：aryl hydrocarbon receptor;G protein-coupled receptor kinase 2;rheumatoid arthritis;CP-25;fibroblasts like synoviocyte;adjuvant-induced arthritis

Foundation item：National Nature Science Foundation of China（81573443;82173824;81973332）;Anhui Province Natural Science Fund（170808J10）;Anhui Provincial Natural Science Foundation（2108085MH320）;and Collaborative Innovation Project of Key Scientific Research Platform in Anhui Universities（GXXT-2020-065）

Corresponding author：CHANG Yan,E-mail：yychang@ahmu.edu.cn;WEI Wei：E-mail：wwei@ahmu.edu.cn

Research progress in pharmacological effects of polyphenols in Areca catechu L.

ZHANG Hao1,PAN Fei-bing2,MA Yun-feng1,ZHANG Dan-shen1,JING Yong-shuai1

（1.College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China;2.Huachuang Institute of Areca Research-Hainan,Haikou 570125,China）

Abstract：Betel nut is the dry and mature seed of Areca catechu L.,which is originated in Malaysia and cultivated in Yunnan,Hainan and Taiwan and other tropical areas of China.It is also known as big belly,binmen,olive seed,green seed and so on.Betel nut is a dual-use resource for medicine and food,which was first contained in LI Dang′s Pharma⁃ceutical Record.Betel nut tastes bitter,pungent,warm in nature,and belongs to the stomach and large intestine meridian.It contains a variety of chemical components such as alkaloids,phenolic compounds,polysaccharides,fatty acids,amino acids,flavonoids,minerals,terpenoids,and steroids.It has the advantages of promoting digestion,lowering blood pres⁃sure,anti-depression,anti-oxidation,anti-inflammatory,and anti-parasites,antibacterial and other activities.The content of total phenols in fresh fruits of areca nut was 31.1%,mainly including catechin,isorhamnetin,chrysopanthoxanthin,luteolin,tannin and other polyphenols.The commonly used methods for determination of polyphenols in areca are vanil⁃lin titration potassium permanganate titration and potassium ferricyanide spectrophotometry.The main activities and mechanisms of areca polyphenols include：①Antidepressant effect：polyphenols bind to monoamine oxidase type A（MAO-A）receptors that inhibit the production of neurotransmitters,thereby increasing the content of amine transmitters in the brain and playing a therapeutic effect on depression.②Antioxidant effect：polyphenols contain multiple adjacent hydroxyl groups,which are easily oxidized and can effectively remove superoxide anion free radical,hydroxyl free radi⁃cal,1,1-diphenyl-2-picrylhydrazyl radical,showing good antioxidant activity.③ Bacteriostatic effect：polyphenols can spe⁃cifically bind to the surface of bacteria,thus achieving bacteriostatic effect.Studies have found that betel nut polyphenols have varying degrees of inhibitory effects on a variety of bacteria.④Inducing apoptosis of lymphocytes：polyphenols deplete the mercaptan in lymphocytes and make them unable to survive,thus inducing apoptosis of lymphocytes.⑤Antiaging effect：polyphenols have the effect of anti-hyaluronidase and anti-elastase,so as to protect elastin fiber and pro⁃mote collagen synthesis.⑥ Anti-allergic effect：studies have found that polyphenols can reduce ovalbumin induced aller⁃gic reactions.⑦Other functions：betel nut can freshen breath,eliminate bad breath,and resist the activity of cobra venom.At present,domestic and foreign scholars′research on betel nut mainly focuses on arecoline and its carcinogenicity,mutagenicity,effects on reproductive function,addiction and toxicity to the nervous system,and there are few studies on the positive effects of betel nut,especially on it.There is less research on phenolic ingredients.Therefore,this article reviews the polyphenolic chemical constituents of betel nut,and fully excavates its pharmacological activity to provide a reasonable basis for the scientific use of betel nut.

Key words：Areca catechu;polyphenols;pharmacological action;scientific utilization

Foundation item：Fund of Dean of Huachuang Institute of Areca Research-Hainan（HCBL2020YZ-012）

Corresponding author：JING Yong-shuai,E-mail：cjys1985@126.com

Research progress in mulberry leaf polysaccharide in treating diabetic nephropathy

ZHANG Shi-lin1,JING Yong-shuai1,ZHANG Dan-shen1,ZHENG Yu-guang2,WU Lan-fang2

（1.College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China;2.College of Pharmacy,Hebei University of Chinese Medicine,Shijiazhuang 050200,China）

Abstract：Diabetic nephropathy（DN）is one of the most common complications of diabetes.It is an important cause of diabetes disability and death.DN is a systemic metabolic syndrome.In its pathogenesis,the interaction of various cell activities and a large number of cytokine biological activities,the activation of signal pathways and so on are involved in the development of DN.At present,the clinical treatment of DN is mainly Western medicine,but it has limitations such as strong toxicity,high side effects and poor compliance.Therefore,the discovery of natural anti-DN substances has also become an important means to treat DN.Mulberry leaves are the dry leaves of Morus alba L.It is not only a tradi⁃tional Chinese medicine,but also a dual-purpose medicinal material for medicine and food.It has the effects of dispelling wind and clearing heat,cooling blood and brightening eyes,tonifying and so on.Mulberry leaf polysaccharide（MLP）is a kind of high molecular compound in mulberry leaves.It has many pharmacological effects,such as hypoglycemic,antiox⁃idant,anti-stress,anti-virus and so on.Therefore,the pharmacological effects of mulberry leaf polysaccharides on dia⁃betic nephropathy are reviewed in this paper,so as to provide references for further research and application.The patho⁃genesis of DN is complex,and the mechanism of renal injury has not been completely clarified.The current studies believe that DN is closely related to heredity,abnormal glucose metabolism,abnormal lipid metabolism,microcirculation disorder,cytokine action,oxidative stress and so on.Relevant studies show that the pharmacological effects of mulberry leaf polysaccharide in the prevention and treatment of DN mainly include：① Effect on transforming factor-β1（TGF-β1）：TGF-β1has become an important cytokine involved in the formation of renal fibrosis by regulating cell proliferation and differentiation and the production of extracellular matrix（ECM）.MLP can significantly inhibit TGF-β1protein,and then inhibit the synthesis of extracellular matrix by renal interstitial fibroblasts and inhibit the realization of fibrosis.②Effect on insulin receptor substrate（IRS-1）：IRS-1 is an important signal molecule at the beginning of IR signal transduction.The decrease of IRS-1 gene expression or the decrease of expression can affect the effective transmission of IR signal and lead to the development and deterioration of diabetes.MPL can significantly increase the expression of IRS-1 mRNA in liver tissue of DN rats,so as to prevent and treat DN.③ Effect on the expression of resistin protein in adipose tis⁃sue.Resistin is a secretory polypeptide derived from adipose tissue and is specifically expressed in white adipose tissue and is closely related to type 2 diabetes mellitus（T2DM）.Experimental studies show that MLP can effectively reduce the expression of resistin protein in white adipose tissue of T2DM rats,indicating that MLP may reduce the level of IR by inhibiting the expression of resistin in adipose tissue,thereby reducing the insulin resistance state of T2DM rats,so as to achieve the goal of treating diabetes.④Effect on adiponectin receptor 1（AdipoR1）：adiponectin can improve insulin resistance,reduce blood glucose and lipid.AdipoR1 is mainly expressed in skeletal muscle and kidney.Studies have shown that AdipoR1 is closely related to the occurrence and development of DN.The results showed that MLP could reduce the blood glucose and blood lipid level and up regulate the expression of AdipoR1 mRNA in DN rats,suggesting that MLP may delay the occurrence and development of DN.This article reviewed the pharmacological effects of mulberry leaf polysaccharides on diabetic nephropathy,and provided a useful basis for further development and utilization of mul⁃berry leaf polysaccharides in the treatment of DN.

Key words：mulberry leaf polysaccharide;diabetic nephropathy;TGF-β1;insulin receptor substrate;resistin;adipo⁃nectin receptor 1

Foundation item：The S&T Program of Hebei Province（H2021423057）

Corresponding author：WU Lan-fang,E-mail：wulanfang757@163.com

Research progress in immune regulation mechanism of Polygonati Rhizoma polysaccharide

ZHANG Ya-meng1,JING Yong-shuai1,ZHANG Dan-shen1,ZHENG Yu-guang2,WU Lan-fang2

（1.College of Chemistry and Pharmaceutical Engineering,Hebei University of Science and Technology,Shijiazhuang 050018,China;2.College of Pharmacy,Hebei University of Chinese Medicine,Shijiazhuang 050200,China）

Abstract：Polygonati Rhizoma is the dry rhizome of Liliaceae plants Polygonatum kingianum coil ethemsl,Polygona⁃tum sibiricum Redoute and Polygonatum cyrtonem Hua.It tastes sweet and has a flat nature.It belongs to the spleen,lung and kidney channels.Polygonati Rhizoma contains a variety of chemical components,including polysaccharides,alkaloids,steroidal saponins,lignans,phytosterols,and so on.Polygonati Rhizoma polysaccharide（PSP）is one of the main bioactive components of Polygonati Rhizoma.It is widely used.It has the effects of enhancing immunity,antiinflammatory,anti-virus and regulating blood lipid.In recent years,the immunomodulatory function of PSP has been paid more and more attention by researchers.PSP can play an immunomodulatory role through a variety of mecha⁃nisms.（1）Effects of PSP on innate immunity.①Macrophages have a strong ability to phagocytize and clear foreign bodies.When polysaccharides bind to macrophage specific membrane receptors,the immune response will be officially activated.RAW264.7 cells can be activated by PSP MR and TLR4 mediated signal pathway to improve the pinocytosis and phagocytosis of RAW264.7 cells.②Natural killer cell（NK cell）is a very important immune cell in the body.It is a non-specific immune killer cell naturally existing in the body.It has the dual functions of immune regulation and cytotoxic⁃ity.It was found that the signal pathway mediated by PSP CR3 and TRL2 may play a major role in the stimulation of NK cells.（2）Effects of PSP on adaptive immune response.①Lymphocytes can be divided into two forms：T cells and B cells due to different differentiation and maturation sites.T lymphocytes are the general name of thymus dependent lym⁃phocytes.B lymphocytes differentiate and mature from animal bone marrow cells and exert their humoral immune func⁃tion by secreting different antibodies.It was found that PSP could activate T/B lymphocytes and increase the ratio of CD4+/CD8+in lymph cells to promote the regulation of immune system.②Thymus and spleen index refers to the level of body immunity through the development of immune organs and the functional status of immune cells.The higher the index of thymus and spleen,the higher the immune activity.A large number of studies have found that PSP can improve immune activity by promoting the proliferation of spleen lymphocytes and regulating organ index,so as to increase the weight and index of thymus and spleen induced by CY.③Antibody is a glycoprotein secreted by B cells after antigen stimulation and a series of proliferation and differentiation into plasma cells.Antibody production level is one of the main indicators of nonspecific immune function.PSP can not only improve the serum antibody level of mice by regulating the phagocytosis of mouse macrophages and the level of serum hemolysin,but also enhance the concentration of IL-2 secreted by spleen lymphocytes in vitro to increase the level of antibody response,and then improve the humoral immune function of the body.（3）Effect of PSP on cytokines.①A large number of experiments have proved that PSP has a significant effect on promoting the production of interleukin（IL）.PSP can combine with specific receptors on the surface of immune cells to activate various intracellular signal transduction pathways,enhance the secretion of cytokines such as IL-2,IL-4,IL-6 and IL-10 by spleen lymphocytes in vitro,make them directly kill target cells and regulate the immune function of the body at the molecular level.②Interferon（IFN）is a special protein or glycoprotein produced by human or animal cells in response to various stimuli.It plays an important role in anti-virus,immune regulation and cell proliferation control.It was found that PSP could increase IFN-γ secreted by T cells and NK cells,activate macrophages to regulate immune function.③Tumor necrosis factor（TNF）is mainly produced by activated macrophages,NK cells and activated T cells.It is a cytokine with important biological activity in antitumor immune response.④Tumor necrosis factor（TNF）is mainly produced by activated macrophages,NK cells and activated T cells.It is a cytokine with important biological activity in antitumor immune response.PSP can promote the proliferation and phagocytic activity of macro⁃phage RAW264.7 to reduce its apoptosis rate.By increasing the secretion of TNF-α,PSP can promote the dissociation between NF-κВ protein and IκВp65 protein after phosphorylation,so as to start the expression and transcription of related immune genes.In conclusion,PSP can improve immunity and has a good application prospect in the development of immunomodulatory drugs.

Key words：polygonatum sibiricum;polysaccharide;immunoregulation;mechanism

Foundation item：The S&T Program of Hebei Province（H2021423057）

Corresponding author：WU Lan-fang,E-mail：wulanfang757@163.com

Ursolic acid ameliorates azoxymethane/dextran sulfate sodium-caused colorectal cancer by inhibition of Wnt signaling cascade

ZHAO Hui1,SUN Qiang1,ZENG Sha1,CHEN Li1,LIU Mao-lun1,REN Shan1,YANG Han1,MING Tian-qi1,TAO Qiu1,LU Jin-jian2,XU Hai-bo1

（1.State Key Laboratory of Southwestern Chinese Medicine Resources,Department of Pharmacology,Chengdu University of Traditional Chinese Medicine,Chengdu 611137,China;2.Institute of Chinese Medical Sciences,University of Macau,Macao,China）

Abstract：OBJECTIVETo investigate the pharmacological effect of ursolic acid（UA）on colitis-associated colorec⁃tal cancer（CAC）and its underlying mechanism based on the Wnt signaling pathway.METHODSThe CAC model in mice was established by azoxymethane（AOM）combined and dextran sulfate sodium salt（DSS）,accompanied by treat⁃ment with various dosages of UA and concomitant appraisal of body weight,stool and physical state of the mice.After the sacrifice of the mice,the tumor and length of the colorectum were measured,followed by retrieval of the liver,spleen,thymus and tumor tissue for downstream assays.The levels of inflammatory factors interleukin-6（IL-6）,IL-1β and C-reactive protein（CRP）in the tumor and serum were examined by enzyme-linked immunosorbent assay（ELISA）.The pathological changes of colorectal tissues were observed by HE staining.The levels in tumors of Wnt/β-catenin sig⁃naling pathway-related proteins Wnt4,GSK-3β,β-catenin,TCF4,LEF1,c-Myc,cyclin D1 and apoptosis-related protein Bcl-2 were assayed by immunohistochemistry（IHC）.The mRNA expressions of Wnt4,GSK-3β,β-catenin,TCF4,LEF1,c-Myc,cyclin D1,Bcl-2,Bax,caspase-9 and caspase-3 in tumors were detected by real-time quantitative RT-PCR（RT-qPCR）.The protein levels of Wnt4,GSK-3β,β-catenin,TCF4,LEF1,c-Myc,cyclin D1,phospho-β-catenin,phospho-GSK-3β,Bcl-2 and Bax in tumors were probed by analyzed by Western blotting（WB）.Also,RNA-seq was employed to assess the gut microbiota in the mice.RESULTSUA significantly ameliorated the symptoms of AOM/DSS-induced mouse CAC,evidenced by improved physical state,body weight,survival rate,colorectal length,the mass of liver,thy⁃mus,spleen,and decreased CAC load and colorectal mass.UA attenuated the levels of IL-6,IL-1β and CRP in the mouse serum and colorectal tumor in a dose-dependent manner.HE staining showed that UA lessened carcinogenesis in the colorectum,with lower infiltration of lymphocytes,versus the control.IHC indicated that UA mitigated the expres⁃sion of Wnt4,β-catenin,TCF4,LEF1,c-Myc,cyclin D1,Bcl-2,and promoted the GSK-3β expression,compared with the control.Furthermore,UA diminished the mRNA expressions of Wnt4,β-catenin,TCF4,LEF1,c-Myc,cyclin D1,Bcl-2,and heightened the mRNA levels of GSK-3β,caspase-3,capase-9 and Bax in CAC.The results of mRNA expressions were verified by WB analysis,which revealed that UA impeded the protein expression of Wnt4,β-catenin,c-Myc,cyclin D1,Bcl-2,TCF4,LEF1,and elevated the protein levels of GSK-3β and Bax,phospho-β-catenin in mouse CAC.In addi⁃tion,UA substantially ameliorated the gut microbiota to store the metabolic function in the mice with CAC.CONCLU⁃SIONUrsolic acid may protect against CAC,potentially by downregulation of Wnt/β-catenin signaling pathway activity and restoration of gut microbiota.

Key words：ursolic acid;colitis associated cancer;Wnt/β-catenin signaling pathway

Foundation item：National Natural Science Foundation of China（81573813;81173598）;Sichuan Provincial Admin⁃istration of Traditional Chinese Medicine of China（2021MS447）;Excellent Talent Program of Chengdu University of Tra⁃ditional Chinese Medicine of China（YXRC2019002,ZRYY1917）;and Open Research Fund of the State Key Laboratory of Southwestern Chinese Medicine Resources of China（2020XSGG006）

Corresponding author：XU Hai-bo,E-mail：xuhb@hotmail.com;LU Jin-jian,E-mail：jinjianlu@um.edu.mo

Hollow-structured nanoparticles as an anticancer drug carrier

ZHU Xiao-ming,WU Xuan,CHEN Jian-li,WAN Hong-ye

（State Key Laboratory of Quality Research in Chinese Medicine,Macau University of Science and Technology,Macao,China）

Abstract：Many drug candidates identified from natural products are poorly water-soluble.The surfactants used to disperse the hydrophobic anticancer drugs in water may cause a serious of acute hypersensitivity reactions.Nanotech⁃nology provides an alternative strategy for delivery of anticancer drugs.Drugs can be encapsulated or attached to the nanomaterials such as lipids,polymers and solid-core nanoparticles.In the present study,porous inorganic nanoparti⁃cles have been utilized for delivery of water-insoluble anticancer drugs.The synthesized nanoparticles were functional⁃ized with different organic polymers.The porous nanoparticles were readily internalized by human glioblastoma U-87 MG cells,and didn′t display cytotoxicity.The internalized nanoparticles were mainly localized in endosomes/lysosomes in cells.With the hydrophobic curcumin and carfilzomib as model drugs,intracellular delivery of hydrophobic anticancer drugs by the porous inorganic nanoparticles was studied.The porous nanoparticle-based encapsulation of hydrophobic drug provides the aqueous dispersion of the drugs.In endosomes/lysosomes mimicking buffers with a pH of 4.5-5.5,pH-dependent drug release was observed from drug loaded nanoparticles.The intracellular drug content and cytotoxicity were significantly higher for drug loaded nanoparticles than free drug.These results suggested porous inorganic nanoparticles might be a promising intracellular carrier for hydrophobic anticancer drugs.

Key words：hollow nanopoarticles;anticancer drug;drug delivery;curcumin

Foundation item：Science and Technology Development Fund,Macau SAR（0168/2019/A3）

Corresponding author：ZHU Xiao-ming,E-mail：xmzhu@must.edu.mo

少突胶质细胞在帕金森病中的研究进展

薄茹雪，陈乃宏，苑玉和

（天然药物活性物质与功能国家重点实验室，中国医学科学院药物研究所，北京 100050）

摘要：少突胶质细胞（OL）是中枢神经系统中高度特化的神经胶质细胞之一，由少突胶质前体细胞（OPC）分化而成。OL通过形成绝缘性髓鞘来促进轴突动作电位的快速传导。帕金森病（PD）是以黑质纹状体多巴胺能神经元丢失为特征的第二大神经退行性疾病，其病理标志物为嗜酸性聚集体路易小体（LB），而LB的主要成分为错误折叠的α-突触核蛋白（α-syn）。研究发现，许多PD患者具有脱髓鞘、白质病变、OL损伤等伴随病理表现，同时，OL中聚集的α-syn能够加速神经元的死亡，提示OL在PD的发生发展过程中发挥着重要的作用。因此，深入研究OL在PD中的作用有助于进一步探索PD的发病机制，有望通过维持髓鞘完整性、保证OL的正常功能而干预PD的病理进展。

关键词：少突胶质细胞；帕金森病；髓鞘病变；α-突触核蛋白

通讯作者：苑玉和，E-mail：yuanyuhe@imm.ac.cn

梓醇通过调节雌激素受体抑制巨噬细胞极化和预防绝经后动脉粥样硬化

陈 琦1＊，张彧涵，张伟薇1＊，毕云慧1，孟庆海1，卞慧敏1，2，李 育1，3

（南京中医药大学1.药学院，2.中药药理与安全性评价江苏省重点实验室，3.医学院与整合医学学院，江苏 南京 210023）

摘要：目的缺乏雌激素会增加绝经后妇女发生动脉粥样硬化（AS）的风险。炎症在AS病理过程中起着至关重要的作用，巨噬细胞与炎症密切相关。梓醇是从传统中药地黄的新鲜根中提取的环烯醚萜苷。本研究旨在评估梓醇对巨噬细胞极化和绝经后AS的影响。方法在体外，用脂多糖（LPS）和干扰素γ（IFN-γ）诱导M1巨噬细胞极化，观察梓醇对巨噬细胞经典活化表型的影响。在体内，用高脂肪饮食喂养ApoE-/-小鼠以诱导AS，并进行卵巢切除术以模拟雌激素缺乏。ig给予梓醇90 d后，取小鼠血液和血管。通过Western印迹、PCR、流式细胞术和免疫荧光等方法观察梓醇对巨噬细胞生物学功能相关标志物和表型转换标志物的影响，以及对雌激素受体表达的影响。同时借助于试剂盒检测梓醇对炎症因子、细胞因子和氧化应激反应相关指标的影响。用ERα和ERβ特异性阻断剂及shRNA干扰雌激素受体表达，以验证梓醇是否通过调节雌激素受体起作用。结果梓醇可以抑制LPS和INF-γ诱导的M1巨噬细胞极化，抑制巨噬细胞M1表型中的促炎反应和氧化应激反应。在体内，梓醇消除绝经后AS小鼠脂质积累，改善血管病变，降低血管钙化程度、斑块面积和纤维帽，明显减缓AS病变进程。梓醇可以上调ERα和ERβ的表达。阻断ERα后，梓醇抗巨噬细胞M1表型的效果降低，抗炎抗氧化应激反应的作用也显著下调，而阻断ERβ对梓醇的抗炎抗氧化应激作用没有显著的影响。结论梓醇可通过增加ERα的表达，抑制巨噬细胞极化，从而预防绝经后AS的发生。

关键词：梓醇；雌激素受体；巨噬细胞；动脉粥样硬化；绝经

通讯作者：卞惠敏，E-mail：hmbian@sina.com；李 育，E-mail：liyu@njucm.edu.cn

＊共同第一作者.

凉膈散抗登革2型病毒的作用

陈 曦，刘俊珊，卢子滨，曹惠慧，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的通过体外实验和动物实验探讨凉膈散（LGS）抗登革2型病毒（DENV-2）的作用。方法体外实验以肝素为阳性药，通过预给药、与病毒联合给药和病毒感染后在给药的方式处理BHK-21细胞，以CCK-8检测细胞活性，Q-PCR和Western印迹法检测E和NS1基因和蛋白的表达，噬斑检测子代病毒释放量评价凉膈散体外抗DENV-2的作用；动物实验以颅内注射加腹腔注射病毒的方法，于ICR乳小鼠上造模，HE染色评价脑和各脏器的病理变化，Q-PCR检测脑、血清、各脏器中E和NS1基因的变化，Western印迹法检测脑E和NS1蛋白的变化。结果体外实验显示，与病毒联合给药时凉膈散具有抗DENV2作用，且高剂量优于阳性药；凉膈散可抑制体内DENV-2，且可以延长感染病毒小鼠的生存时间以及改善临床症状。结论凉膈散具有抗DENV-2作用。

关键词：凉膈散；登革病毒；抗病毒；E；NS1

通讯作者：余林中，E-mail：yulzh@smu.edu.com

增溶型白桦脂酸纳米制剂的制备及其在体内外抑制乳腺癌生长的作用及机制

陈泽坤，郭传龙

（青岛科技大学，山东 青岛 266042）

摘要：目的制备增溶型白桦脂酸（BA）纳米制剂，并探讨其在体内外抑制乳腺癌生长的作用及机制。方法以Soluplus为载体，采用薄膜水化法制备包载BA的纳米制剂（Soluplus-BA）并进行表征；MTT法检测Soluplus-BA制剂和游离BA对乳腺癌细胞MDA-MB-231增殖的影响；DCFH-DA和JC-1染色检测Soluplus-BA对细胞活性氧和线粒体膜电位的影响；γ-H2AX免疫荧光染色检测Soluplus-BA对细胞DNA损伤的影响；HUVEC细胞成管实验和鸡绒毛尿囊膜（CAM）实验检测Soluplus-BA是否可抑制血管新生；Western印迹法检测相关信号通路蛋白的表达；小鼠移植瘤实验检测Soluplus-BA在体内的抗肿瘤活性。结果制备的Soluplus-BA制剂的平均粒径为（54.77±1.26）nm，多分散指数（PDI）为0.083，并在9个月内存储稳定性好。MTT测定结果显示，Soluplus-BA制剂增强了BA对MDA-MB-231细胞生长的抑制作用，IC50分别为（38.81±4.9）mg·L-1（Soluplus-BA）和（15.45±3.01）mg·L-1（BA）。Soluplus-BA制剂可通过诱导MDA-MB-231细胞活性氧的积累和线粒体膜电位的破坏发挥抗肿瘤作用。免疫荧光结果显示γH2AX荧光斑点的增加，表明Soluplus-BA制剂诱导了DNA双链断裂。Soluplus-BA制剂还抑制人脐静脉内皮细胞（HUVEC）的小管形成和迁移，并抑制鸡绒毛尿囊膜的新生血管形成。Western印迹法结果显示，Soluplus-BA通过抑制HIF-1/VEGF-FAK信号通路发挥抑制血管新生活性。小鼠体内结果显示，Soluplus-BA制剂显著抑制乳腺癌移植瘤的生长，抑瘤率高于游离BA。结论增溶型Soluplus-BA胶囊增加了BA在体内外的抗肿瘤活性，并调控HIF-1/VEGF-FAK信号通路。

关键词：白桦脂酸；纳米制剂；抗乳腺癌

基金项目：山东省自然科学基金（ZR2020QH359）

通讯作者：郭传龙，E-mail：guochuanlong@qust.edu.cn

双氢青蒿素通过增强免疫共抑制信号改善小鼠实验性自身免疫性脑脊髓炎

杜欣珂，冉庆森，李 琦，刘 丽，孙立东，杨 庆，李玉洁，陈 颖，翁小刚，王娅杰，蔡维艳，朱晓新

（中国中医科学院中药研究所，北京 100700）

摘要：目的探究双氢青蒿素（DHA）对小鼠实验性自身免疫性脑脊髓炎模型（EAE）的改善作用及其作用机制。方法使用髓少突细胞糖蛋白（MOG35−55）诱导雌性C57BL/6小鼠神经炎症和髓鞘脱失，并重点分析急性期内给药组和模型组的生理功能和组织病理结构。通过流式细胞术检测小鼠脾细胞免疫应答功能。Western印迹法和qRT-PCR分别检测相关蛋白和基因表达水平。结果与EAE模型小鼠相比，用DHA治疗的小鼠脊髓损伤更小，炎症细胞浸润更少。从机制层面看，DHA增强了脾细胞中细胞毒性淋巴细胞抗原4（CTLA4）和程序性细胞死亡1（PD1）的表达。此外，DHA上调细胞因子信号抑制因子3（SOCS3）的表达和转录信号转导激活因子1（STAT1）的磷酸化。结论DHA可通过STAT1/SOCS3途径，上调T细胞CTLA4和PD1的表达，改善EAE模型小鼠病理症状。本研究提示出DHA作为多发性硬化症治疗药物具有巨大的开发潜力。

关键词：双氢青蒿素；实验性自身免疫性脑脊髓炎；细胞毒性淋巴细胞抗原4；程序性细胞死亡1

基金项目：国家科技重大专项（2017ZX09101002-002-002）；中国中医科学院优秀青年科技人才专项培养计划（ZZ13-YQ-044）；中国中医科学院“一带一路”合作项目〔（20141202）GH201914〕；中药研究所内部项目（Z2017019-03）；国家自然科学基金（8157141621）

通讯作者：朱晓新，E-mail：zhuxx@icmm.ac.cn

白头翁皂苷A3通过抑制巨噬细胞M2型极化增强结肠癌细胞对5-氟尿嘧的化疗敏感性

范泽萍1，2，陈兰英1，2，刘 鹏1，2，崔亚茹1，2，罗颖颖1，2

〔1.江西中医药大学中药固体制剂制造技术国家工程研究中心，江西南昌330006；2.中药药效（扶正祛邪抗肿瘤）评价重点研究室，江西南昌 330006〕

摘要：目的探讨白头翁皂苷A3通过调控巨噬细胞极化对5-氟尿嘧啶（5-FU）化疗敏感性的影响及其作用机制。方法利用白细胞介素（IL）-4和IL-13诱导巨噬细胞M2极化，采用CCK-8法检测白头翁皂苷A3对巨噬细胞增殖的影响；采用流式细胞术检测白头翁皂苷A3对巨噬细胞极化表型的影响；采用RT-PCR检测白头翁皂苷A3对巨噬细胞CD206，IL-10，CCL18，CCL22 mRNA表达的影响。收集M2巨噬细胞及白头翁皂苷A3干预M2巨噬细胞后的上清液作为条件培养基（CM）与5-FU共同作用于结肠癌SW480细胞。采用CCK-8法检测CM干预下5-FU对SW480细胞增殖的影响；采用Annexin-FITC/PI双染法及Western印迹法检测CM干预下5-FU对SW480细胞凋亡及凋亡蛋白Bax、活化的胱天蛋白酶3表达的影响。结果白头翁皂苷A3浓度＜100 mg·L-1时对巨噬细胞增殖无抑制作用，故选择50，75和100 mg·L-1作为白头翁皂苷A3实验浓度。白头翁皂苷A3呈浓度依赖性降低CD206的阳性细胞表达比例，显著抑制M2巨噬细胞表面抗原CD206和极化因子IL-10，CCL18，CCL22 mRNA的表达（P＜0.01）。白头翁皂苷A3 100 mg·L-1能明显提高了5-FU对SW480细胞的抑制率（P＜0.05），同时提高了SW480细胞的凋亡率及促凋亡蛋白Bax和活化的胱天蛋白酶3的表达（P＜0.05）。结论白头翁皂苷A3通过抑制巨噬细胞M2极化减弱了M2巨噬细胞对SW480细胞促凋亡蛋白Bax和活化的胱天蛋白酶3的抑制作用，从而降低M2巨噬细胞介导的SW480细胞对5-FU的化疗抵抗，提高了5-FU的化疗敏感性。

关键词：白头翁皂苷A3；巨噬细胞极化；结肠癌SW480细胞；5-氟尿嘧啶；化疗敏感性

基金项目：国家自然科学基金（81860720）；江西省中医药重点研究室建设项目

通讯作者：陈兰英，E-mail：clyxy2513@163.com

藏药仁青常觉对脂多糖诱导大鼠肺泡Ⅱ型上皮细胞损伤的保护作用

高小敏，姜 一，孔秀梅，赵 勤

（西藏民族大学医学院，西藏民族大学教育部藏药检测工程中心，陕西 咸阳 712082）

摘要：目的利用体外实验研究藏药仁青常觉对脂多糖（LPS）诱导的大鼠肺泡Ⅱ型上皮细胞（RLE-6TN）损伤的保护作用。方法用不同浓度的水提仁青常觉干预RLE-6TN 2 h，再加入LPS 1 mg·L-1刺激细胞24 h造成炎症损伤，收集细胞上清，用ELISA测定白细胞介素6（IL-6）、肿瘤坏死因子α（TNF-α）和IL-10的水平，并检测细胞Bcl-2、Bax和胱天蛋白酶3蛋白表达水平。结果水提仁青常觉可明显降低LPS诱导的RLE-6TN的IL-6和TNF-α高表达，显著升高IL-10的表达，升高细胞中Bcl-2/Bax蛋白表达水平，降低胱天蛋白酶3的蛋白表达。结论仁青常觉对LPS诱导的RLE-6TN损伤具有保护作用，其作用机制可能与降低炎症因子的释放和抗凋亡有关。

关键词：仁青常觉；脂多糖；大鼠肺泡Ⅱ型上皮细胞；白细胞介素6；白细胞介素10；肿瘤坏死因子α

基金项目：西藏自治区自然科学基金重点项目（XZ202101ZR0076G）；西藏民族大学新冠肺炎疫情应急重点项目（XZM⁃DYJ02）

通讯作者：赵 勤，E-mail：xyzhaoqin@126.com

八味沉香丸对垂体后叶素致大鼠急性心肌缺血的保护作用

韩艺玮，孔秀梅，姜 一，张晓英

（西藏民族大学医学院，西藏民族大学藏药检测技术教育部工程研究中心，西藏藏医药研究中心联合实验室，陕西 咸阳 712082）

摘要：目的通过舌下注射垂体后叶素（PIT）建立急性心肌缺血模型，观察八味沉香丸对心肌缺血的保护作用。方法采用大鼠舌下静脉注射PIT制急性心肌缺血模型，以0.03 g·kg-1剂量的复方丹参片为阳性对照，观察大鼠心电图T波和ST段的改变、检测血清中乳酸脱氢酶（LDH）和肌酸激酶（CK）的活性以及采用HE染色观察心肌组织病理形态来评价八味沉香丸对心肌损伤的保护作用。结果与对照组相比，垂体后叶素诱导的心肌缺血大鼠心电图T波和S-T段明显升高，血清中心肌酶CK和LDH的释放量增加，HE染色观察到心肌细胞排列紊乱，血管充血明显。八味沉香丸0.5和1.0 g·kg-1ig给药10 d后，心电图T波和ST段（反映心室的复极过程）的变化值较模型组明显降低。HE染色发现，药物组心肌组织的血管充血明显减轻，心肌细胞排列整齐，横纹清晰。同时检测到大鼠血清LDH和CK的释放量减少（P＜0.01）。结论八味沉香丸能显著改善心肌缺血大鼠S-T段和T波的变化，增加心肌收缩力，减轻心肌缺血引起的炎症反应，降低血清中LDH和CK的活性，对急性心肌缺血损伤具有一定的保护作用。

关键词：八味沉香丸；垂体后叶素；心肌缺血；乳酸脱氢酶；肌酸激酶

项目基金：西藏自治区自然科学基金［XZ2018ZR G-85（Z）］；陕西省教育厅专项科学研究计划（19JK0890）

NLRP3炎症小体在机体脂质代谢中的作用研究进展

贺润铖，黄芷棋，冯倩倩，董世芬

（北京中医药大学中药学院中药药理系，北京 102488）

摘要：在世界范围内，肥胖人口正逐年增加并呈现年轻化趋势，伴随增加了许多疾病风险如脂肪肝、2型糖尿病等。在肥胖患者的机体中，脂肪组织作为内分泌组织分泌脂肪因子，并促进巨噬细胞及其他免疫细胞的募集和浸润，释放炎症细胞因子和趋化因子，最终导致炎症并影响机体代谢。核苷酸结合寡聚化结构域样受体蛋白3（NLRP3）炎症小体是一种蛋白复合体，胞质内由多种蛋白构成，可通过激活胱天蛋白酶1，释放IL-1β和IL-18等炎症因子，发挥炎症效应。机体脂质代谢紊乱与NLRP3炎症小体密切相关，在肥胖者体内存在着内源性危险信号如过量的ATP、葡萄糖和饱和脂肪酸等，能激活NLRP3炎症小体，促使胱天蛋白酶1介导的加工、分泌IL-1β和IL-18等炎症因子，最终导致脂肪组织炎症、破坏脂肪分解，引起代谢功能紊乱。NLRP3炎症小体的激活还会干扰机体内白色脂肪褐变以及促进棕色脂肪“白化”，有研究表明，NLRP3炎症小体分泌的IL-1β可干扰β3-R激动剂诱导的白色脂肪细胞褐变，降低脂肪细胞产热基因如UCP1和PCG-1表达，阻断NLRP3基因表达或阻断IL-1β分泌可改善这种情况。NLRP3炎症小体不但直接参与肥胖相关的脂肪组织炎症和功能失调，而且可以通过影响包括钙敏感受体、血清网膜素-1和沉默信息调节因子1等多种肥胖相关因子调控该过程。因此进一步探究NLRP3炎症小体在脂质代谢中的作用机制对肥胖及其引起的相关疾病提供了新思路。

关键词：NLRP3炎症小体；脂质代谢；肥胖；炎症因子

基金项目：国家自然科学基金（81503287）

通讯作者：董世芬，E-mail：dongshifen@bucm.edu.cn

联合网络药理学分析黄芪葛根汤抗阿尔茨海默病治疗作用

侯雯倩1，苏 敬1，刘小凤1，董劲曲1，徐 兰1，海 洋2，刘东玲1

（甘肃中医药大学1.药学院，2.科研实验中心，甘肃 兰州 730000）

摘要：目的阿尔茨海默病（AD）是世界第一的神经退行性疾病，目前缺乏有效的治疗药物。研究发现，AD与2型糖尿病（T2DM）之间存在密切联系。课题组系统分析了19种糖尿病治疗复方，从中筛选出具有AD治疗作用的黄芪葛根汤，并通过网络药理学以及现代药理学基础分析探讨黄芪葛根汤治疗AD的潜在靶点及信号通路，阐述其可能的作用机制。方法采用Aβ1-42基因诱导的秀丽隐杆线虫作为AD病理模型，探讨黄芪葛根汤的抗AD作用。从TCMSP和BATMAN-TCM数据库搜索黄芪葛根汤中2味中药的化合物及靶点，使用GeneCards、NCBI、CTD以及OMIM数据库检索AD和T2DM的靶点；通过STRING数据库构建蛋白质相互作用网络；通过cytoHubba，选择MCC对关键靶点进行分析；采用Metascape数据库对关键靶点进行聚类分析；采用Omicshare平台对关键靶点进行KEGG分析及GO分析，以获取潜在靶标与机制。结果黄芪葛根汤共包含2味中药，14种化合物，相对应的药物靶标有113个，黄芪葛根汤、AD、T2DM共同交集靶点共31个。对交集靶点进行MCC聚类分析获得JUN，PTGS2和SIRT1等核心靶点。交集靶点可以聚类为3类，分别与过氧化物酶体增殖剂激活受体信号传导途径、发育生长和凋亡信号通路有关，这些途径和AD密切相关。通过KEGG信号通路富集筛选所得排名前三的信号通路分别为钙信号通路、神经活性配体-受体相互作用和cAMP信号通路。GO分析得到主要的生物过程聚类分析包括氮化合物的细胞反应、血液循环和神经递质水平的调节；细胞组成聚类分析主要有突触后膜、突触后、突触膜；核心分子功能聚类分析包括神经递质受体活性、核糖核酸聚合酶Ⅱ特异性脱氧核糖核酸结合转录因子结合，以及乙酰胆碱结合。另外，秀丽隐杆线虫实验结果表明，黄芪葛根汤可以延缓线虫肌肉组织因过度表达Aβ1-42导致麻痹表型的AD样病理特征，其中0.5 g·L-1的抗AD活性最优。进一步实验证明黄芪葛根汤还可显著减缓泛神经元过度表达Aβ1-42导致的线虫对外源5-羟色胺超敏感的AD样病理特征。结论黄芪葛根汤可能通过对钙离子稳态失衡、线粒体氧化应激、炎症等的调节治疗AD，且实验研究证明黄芪葛根汤对AD线虫有治疗作用。

关键词：黄芪葛根汤；阿尔茨海默病；网络药理学；秀丽隐杆线虫

基金项目：甘肃省高等学校创新基金（2021B-160）；甘肃省中医药研究中心开放课题（zyzx-2020-21）；甘肃中医药大学科学研究与创新基金（2020KCYB-1）

通讯作者：刘东玲，E-mail：dongling83@163.com；海洋，E-mail：cathyhaiyang@163.com

酸枣仁中黄酮类物质药理作用研究进展

胡金颖1，代立霞2，张丹参1，景永帅1

（1.河北科技大学化学与制药工程学院，河北 石家庄 050018；2.石家庄市中医院肛肠科，河北 石家庄 050051）

摘要：中药酸枣仁是鼠李科植物酸枣〔Ziziphous jujuba Mill.Var.spinosa（Bunge）Hu.ex H.F.Chou〕的干燥成熟种子，性平，味甘、酸，具有补肝、宁心、敛汗、生津之功，是最早被列为药食同源的中药之一，酸枣仁化学成分主要包括酸枣仁皂苷、酸枣仁黄酮、酸枣仁油等。现代药理研究表明，黄酮类化合物是其主要的活性成之一，黄酮类化合物主要活性成分有斯皮诺素及6′-阿魏酰斯皮诺素，具有镇静催眠、抗抑郁、抗焦虑、抗氧化以及提高记忆力等功效的作用。①镇静催眠作用：6-羟基黄酮与γ-氨基丁酸（GABA）受体/苯二氮受体有很强的亲和力，通过影响GABA的含量的方式来影响睡眠；斯皮诺素显著延长由戊巴比妥所诱导的睡眠时间，缩短睡眠潜伏期，并成剂量依赖性，研究发现其主要是通过5-羟色胺（5-HT）的调节降低小鼠脑组织内多巴胺和盐酸去甲肾上腺素（NE）含量来发挥镇静催眠作用。②抗焦虑作用：斯皮诺素通过降低海马中NE的释放，抑制海马中5-HT的合成，降低血浆β-内啡肽含量，提高血清一氧化氮浓度来发挥抗焦虑作用，表明其作用机制与增加小鼠脑内GABA量、降低N-甲基-D-天冬氨酸（NMDA）受体1含量和谷氨酸表达有关。③抗抑郁作用：酸枣仁黄酮类物质通过调控血浆糖皮质激素，进而调控NMDA受体中NR1和NR2A/2B的表达来发挥抗抑郁作用。④抗氧化作用：酸枣仁总黄酮可改善D-半乳糖诱导的氧化损伤大鼠的记忆损伤及氧化应激作用，实验结果表明其主要通过调节牛磺酸和次牛磺酸代谢，甘氨酸、丝氨酸和苏氨酸代谢以及丙酮酸代谢发挥抗氧化损伤作用。⑤增强学习记忆的作用：斯皮诺素显著增加了海马体中磷酸化细胞外信号调节激酶和环腺苷酸结合蛋白酶的表达水平，从而起到增强学习记忆的作用。⑥对脑神经保护作用：酸枣仁总黄酮通过激活下丘脑室旁核cAMP-PKA-CREB信号通路，抑制脑组织c-fos基因表达和神经细胞凋亡，从而发挥脑神经保护作用。酸枣仁黄酮的药理学作用广泛多样，药理作用机制复杂，对于酸枣仁黄酮单体的研究，着重在于斯皮诺素的研究，其更加深入的药理作用及其他单体成分药理活性研究仍是有待研究的热点。本文对酸枣仁中的黄酮类化学成分的药理作用进行系统性总结，以期为酸枣仁的进一步的研究和临床应用提供参考。

关键词：酸枣仁；黄酮类；镇静催眠；抗抑郁；抗焦虑

基金项目：河北省中医药管理局中医药科研计划（2020331）

通讯作者：景永帅，E-mail：cjys1985@126.com

黄连解毒汤对阿尔茨海默病模型斑马鱼认知功能障碍的影响

黄 瑶，刘凯菲，关国凯，陈毅飞

（桂林医学院药学院，广西 桂林 541199）

摘要：目的探究黄连解毒汤对阿尔茨默病（AD）斑马鱼认知功能障碍的影响。方法将126 g黄连，84 g黄柏，84 g黄芩，126 g栀子分别放入10，8和8倍量的水，浸泡30 min，然后100℃分别提取2，1.5和1.5 h，将3次提取液粗滤浓缩为308 g·L-1放入冰箱备用。选取200尾斑马鱼于第1～7天在T型缸进行行为学训练，结束后录像并利用行为学记录分析软件smart3.0进行行为学分析，选出合格斑马鱼，为空白组。除空白组斑马鱼不做任何处理外，其余5组于第8～13天暴露在AlCl3100 μg·L-1养殖水中进行造模，结束后录像并利用行为学记录分析软件Smart3.0进行行为学分析，选出造模成功的斑马鱼，之后于第14～19天分别给药石杉碱甲4 μg·L-1和黄连解毒汤（154，308和616 mg·L-1），结束后录像并通过行为学记录分析软件smart3.0进行行为学分析，选出合格的斑马鱼。训练结束后每组斑马鱼取脑，通过Western印迹法和qPCR检测N-cadherin、p-p38 MAPK/p38 MAPK和p-tau的表达变化来表现黄连解毒汤对AD斑马鱼认知功能障碍的影响。结果Western印迹和qPCR结果显示，给予黄连解毒汤后N-cadherin含量较模型组增加（P＜0.05），p-p38 MAPK/p38 MAPK和p-tau的含量较模型组降低（P＜0.05）。结论黄连解毒汤对AD模型斑马鱼认知功能障碍有一定的缓解作用。

关键词：黄连解毒汤；阿尔茨海默病；斑马鱼；认知功能障碍

基金项目：广西壮族自治区自然科学基金（2017GXNSFAA198255；2018GXNSFBA138028）；广西脑与认知神经科学重点实验室开放课题（GKLBCN-20180105-03）；2019大学生创新创业项目训练计划（201910601038）；第四期广西高等学校千名中青年骨干教师培育计划

通讯作者：陈毅飞，E-mail：chenyifei_pharm@glmc.edu.cn

痰热清注射液抑制登革2型病毒诱导的小胶质细胞炎症反应

黄鹤飞，刘俊珊，曹慧慧，卢子滨，余林中

（南方医科大学中医药学院中药药理学教研室，广东 广州 510515）

摘要：目的研究痰热清注射液对登革2型病毒（DENV-2）感染的小鼠小胶质细胞（BV2）炎症细胞因子的影响。方法BV2细胞铺板后分组感染并给药，分为5组，空白组，DENV-2感染组，痰热清注射液低、中和高剂量组。培养24 h后收取上清液和细胞，实时RT-PCR分别检测BV2的IL-6、TNF-α mRNA表达水平；双抗夹心ELISA法检测培养上清中上述细胞因子表达。以颅内注射的方法于ICR乳小鼠上造模，苏木素-伊红（HE）染色评价脑和各脏器的病理变化。结果体外实验显示，与病毒联合给药时痰热清具有抗DENV-2作用，可降低炎症因子水平；进一步在体内实验表明，痰热清注射液可以延长感染病毒小鼠的生存时间以及改善症状。结论痰热清注射液可用于缓解登革2型病毒感染引起的脑炎。

关键词：登革病毒；痰热清；炎症；小胶质细胞

通讯作者：余林中，E-mail：yulzh@smu.edu.cn

基于网络药理学和分子对接探讨藏药麻花秦艽-沙棘配伍抗缺氧作用及机制

姜 一，孔秀梅，韩艺玮，赵 勤

（西藏民族大学医学院教育部藏药检测工程中心，陕西 咸阳 712082）

摘要：目的通过网络药理学的方法对中药沙棘和藏药麻花秦艽配伍防治缺氧的主要活性成分和作用机制进行预测和分析，为临床用药和开发新药提供依据。方法通过中国知网、万方等数据库、中药系统药理学数据库分析平台（TCMSP）和Swiss Target Prediction平台，收集沙棘和麻花秦艽的活性化合物并对其靶点进行预测，应用人类基因数据库（GeneCard）收集缺氧的相关靶点；用Cytoscape软件来构建沙棘，麻花秦艽活性成分和对应的靶点的成分-靶点网络图；利用STRING数据库构建药物疾病交集靶点的PPI网络，并筛选出关键靶点和中心靶点；利用David数据库进行Go分析和KEGG信号通路富集分析，利用AutoDock软件进行分子对接，利用Pymol软件进行相应的制图。结果共筛选出沙棘和麻花秦艽活性成分35个（如熊果酸、山奈酚、木犀草素等），与高原缺氧相关的靶点162个，其中关键靶点28个（如PIK3R1，MAPK3，IL-2和MAPK等）。GO功能分析确定了107个条目（P＜0.05），涉及凋亡的正负调控作用和氧化应激反应等。KEGG通路分析发现相关的信号通路39条（如Rap1信号通路、ErbB信号通路、肾素-血管紧张素系统等），分子对接结果显示，多数活性成分与关键靶点具有结合作用。结论本研究初步证明了麻花秦艽和沙棘配伍对缺氧性疾病的预防和治疗有着多靶点多通路的特点。

关键词：麻花秦艽-沙棘；网络药理；缺氧；靶点；信号通路；分子对接

基金项目：国家自然科学基金（81660722）；西藏民族大学重点项目（13myZP07）

通讯作者：赵 勤，E-mail：xyzhaoqin@126.com

真菌次级代谢产物Rasfonin对鸟苷酸交换因子SOS1表达的调控作用及机制

李 静1，2，3，肖智勇1，2，周文霞1，2，张永祥1，2

（1.军事科学院军事医学研究院毒物药物研究所，北京 100850；2.抗毒药物与毒理学国家重点实验室，北京 100850；3.南京中医药大学，江苏 南京 210023）

摘要：目的Rasfonin是一种从青藏高原海拔＞4 km土壤样品的真菌中发酵提取的α-吡喃酮类化合物。前期研究表明，Rasfonin可下调SOS1（Son of sevenless）蛋白表达以及由其介导的RAS蛋白核酸交换反应，但作用机理尚不明确。本研究拟探讨Rasfonin对SOS1表达的调控以及KRAS蛋白在其中的参与作用，以期进一步阐述Rasfonin的抑癌机制，为抗肿瘤新药的研究提供实验依据。方法在野生型KRASWT、突变型KRASG12C和KRASG12D的人源肿瘤细胞上观察Rasfonin对SOS1表达的作用，应用Western印迹法和实时荧光定量PCR检测SOS1蛋白和mRNA表达；采用双荧光素酶报告基因实验测定SOS1启动子活性；在此基础上，将KRASWT、KRASG12D、KRASG12C和SOS1质粒转染293T细胞构建KRAS与SOS1共表达体系来探讨KRAS在Rasfonin抑制SOS1表达过程中的作用。结果Rasfonin的阴性对照化合物A321不影响SOS1蛋白表达。Rasfonin作用后，KRAS野生型细胞SOS1蛋白表达量不变，mRNA短暂升高后恢复至对照水平，而突变型细胞SOS1蛋白表达量明显减少（P＜0.05），mRNA短暂升高后显著降低（P＜0.05）。293T细胞中无KRAS蛋白时，Rasfonin对SOS1启动子活性无影响，而转染KRAS（WT，G12C和G12D）质粒后，Rasfonin明显抑制SOS1启动子活性（P＜0.05）。单转染SOS1及其与野生型KRAS共转染时，Rasfonin仅轻微下调SOS1蛋白表达；而共转染SOS1与突变型KRAS（G12C和G12D）时，Rasfonin可显著抑制SOS1蛋白表达（P＜0.05）。结论Rasfonin可能通过突变型KRAS（G12C和G12D）蛋白抑制SOS1的启动子活性，进而选择性下调突变型KRASG12C和KRASG12D肿瘤细胞中SOS1 mRNA及蛋白的表达水平。本课题深化了对Rasfonin抑癌机制及SOS1表达调控的认识，为RAS突变肿瘤的治疗提供了新的思路。

关键词：Rasfonin；SOS1；KRAS；表达调控；胰腺癌

项目基金：国家自然科学基金（81803431）

通讯作者：肖智勇，E-mail：zy-xiao@163.com；周文霞，E-mail：zhouwx@bmi.ac.cn

马钱子碱通过钾离子通道调节外周镇痛

李永丰

（陕西中医药大学陕西省针药结合重点实验室，陕西 咸阳 712046）

摘要：目的马钱子是传统的镇痛中药之一，马钱子碱（又名布鲁生）是马钱子镇痛的有效成分，但外周镇痛机制研究较少，本文通过痛行为检测和单纤维自发放电手段，观察马钱子碱在外周损伤区中的镇痛作用机制。方法建立慢性压迫损伤模型（CCI），利用机械缩足反射阈值（PWT）和热缩足潜伏期（PWL）检测大鼠的疼痛行为学变化；记录起步点单纤维自发放电检测马钱子碱对放电频率的影响。结果与溶剂组相比，外周损伤区局部注射马钱子碱后PWT和PWL显著增加（P＜0.01），预先注射非特异性钾通道阻断剂四乙基铵（TEA）/大电导钙激活钾离子通道（BKca）阻断了马钱子碱的镇痛作用；马钱子碱使起步点自发放电频率显著降低（P＜0.01），而灌流TEA后抑制了它的作用，进一步灌流钙依赖钾离子的特异性阻断剂阻断BKca、中电导钙激活钾离子通道和小电导钙激活钾离子通道，发现仅阻断BKca通道可抑制马钱子碱对起步点自发放电频率的抑制作用。结论马钱子碱在外周具有镇痛作用，并且BKca能够阻断其作用，表明BKca参与马钱子碱在外周的镇痛作用。

关键词：慢性压迫损伤模型；马钱子碱；大电导钙激活钾离子通道；镇痛

八角枫碱的神经毒性及机制

练东银，侯红平，张广平，李建荣，叶祖光，彭 博

（中国中医科学院中药研究所，北京 100700）

摘要：目的采用人神经母细胞瘤SH-SY5Y和人神经胶质瘤U251细胞作为体外评价模型，初步探讨八角枫药材中的主要活性成分八角枫碱的神经毒性及其机制。方法采用CCK-8和中性红摄取实验测定细胞毒性；采用Hoechst 33342核酸染色法检测染色质凝集情况；分光光度法检测胱天蛋白酶3和9的表达水平；Western印迹法检测胶质纤维酸性蛋白（GFAP）、波形蛋白、Bax、PARP和ERK信号通路蛋白的表达及磷酸化水平；采用维甲酸诱导神经元分化，检测分化的SH-SY5Y细胞神经突生长和退化。结果与对照组相比，八角枫碱50～2000 mg·L-1能明显抑制U251和SH-SY5Y的细胞生长（P＜0.05，P＜0.01），作用呈浓度和时间依赖性，24 h的 IC50值分别为（770.83±29.95）mg·L-1（U251）和（668.05±9.75）mg·L-1（SH-SY5Y）。八角枫碱400 mg·L-1作用于U251细胞24 h后，显著抑制胶质细胞标志物GFAP和波形蛋白蛋白的表达。八角枫碱200，400，600和800 mg·L-1处理24 h后，U251细胞和SH-SY5Y细胞出现明显凋亡形态学改变，表现为细胞核或细胞质内可见浓染致密的颗粒块状荧光；与对照组相比，八角枫碱能剂量依赖性明显增加胱天蛋白酶3和9的酶活性（P＜0.05，P＜0.01），其中U251细胞内胱天蛋白酶9的酶活力值（U）为7.76±0.06，8.61±0.89，9.23±0.34和7.48±0.30；SH-SY5Y细胞内胱天蛋白酶9的酶活力值（U）为2.15±0.14，2.38±0.45，4.22±0.59和3.15±0.75，胱天蛋白酶3的酶活力值（U）为0.89±0.06，0.96±0.04，1.27±0.32和1.1±0.29。Western印迹法结果显示，八角枫碱能上调Bax表达，促进PARP切割，明显抑制ERK蛋白的磷酸化水平。八角枫碱50，100，200和300 mg·L-1作用于分化的SH-SY5Y细胞24 h后，神经突长度（μm）为80.87±21.37，64.65±14.46，46.58±12.36和15.32±4.61，相对于对照组（87.54±21.68）μm，神经突长度变短（P＜0.05，P＜0.01），表明八角枫碱可剂量依赖性抑制分化的SH-SY5Y神经突的生长及诱导其神经突退化。结论八角枫碱能抑制神经细胞及神经胶质细胞的增殖，并抑制神经元轴突的生长，其作用机制可能通过抑制MAPK-ERK通路，上调Bax表达，促进PARP切割，增加胱天蛋白酶3和9活性，从而促进线粒体依赖性细胞凋亡。

关键词：八角枫碱；神经毒性；MAPK-ERK信号通路

基金项目：国家重点研发计划（2018YFC1708105）

通讯作者：彭 博，E-mail：bpeng@icmm.ac.cn

基于HPLC-MS/MS法的补肾益智方君药蛇床子素抗阿尔茨海默病药动学

梁 勇1，2，刘金满1，2，伍帆昌1，2，黄明俊1，2，王 奇1，2

（广州中医药大学1.科技创新中心，2.临床药理研究所，广东 广州 510405）

摘要：目的考察补肾益智方君药蛇床子素的药动学，探究复方增强蛇床子素生物利用度的机制。方法通过构建HPLCMS/MS高灵敏度的蛇床子素准确定量分析方法，以多反应检测（MRM）方式，以欧前胡素为内标，建立蛇床子和欧前胡素的母离子（Q1）和子离子（Q3）的离子对；进一步优化HPLC-MS/MS参数，实现蛇床子素的准确定量分析。以SD大鼠为模型动物，通过ig给药方式，设置对照组、假手术组和手术组，用连续给药的方式，眼眶采血，时间点为0.5，1，2，4，6，8，10，12和24 h，基于HPLC-MS/MS测定蛇床子素的含量。结果给药4 h后，大鼠血浆中蛇床子素浓度达到峰值，为2426 μg·L-1，药效持续时间为6 h。质谱方法检出限为1.8 μg·L-1，线性系数0.9998，准确度为94.6%～105.2%。结论补肾益智方较之蛇床子素可显著提高蛇床子素的药动学，且基于HPLC-MS/MS的蛇床子素分析方法具有灵敏度高、准确性好的特点，为进一步研究蛇床子素抗AD机制提供了强大的分析检测方法。

关键词：阿尔茨海默病；补肾益智方；蛇床子素；HPLC-MS/MS；药动学

六味地黄汤活性成分的抗补体作用

林 颖1，2，3，韩 露1，2，肖智勇1，2，周文霞1，2

（1.军事科学院军事医学研究院毒物药物研究所，北京 100850；2.抗毒药物与毒理学国家重点实验室，北京 100850；3.南京中医药大学，江苏 南京 210023）

摘要：目的补体系统是人体重要的免疫防御系统之一，类风湿性关节炎、老年痴呆、系统性红斑狼疮等多种疾病中存在补体异常激活的现象，对人类健康造成极大的威胁。中药成分如黄酮、多糖、蒽醌、甾体和生物碱等具有广泛的抗补激活的作用，因此从中筛选抗补体活性成分具有较好的应用前景。本研究探讨六味地黄汤活性成分的抗审体作用。方法通过采用对自主采集和公开的108种中药复方及其中的单体成分作用于细胞的转录组数据进行基于KEGG通路的富集分析，预测具有潜在抗补体活性的成分，然后通过采用体外细胞溶血法对以上计算获得的中药活性部位进行抗补体活性验证。结果转录组分析数据显示，六味地黄汤的活性成分糖苷提取物D-b的下调基因显著富集于补体级联激活通路（P＜0.01），补体经典途经溶血实验显示，D-b具有呈浓度依赖性抑制红细胞溶血的作用，D-b对红细胞的溶血抑制率略弱于肝素，其中100 mg·L-1浓度的D-b和肝素的溶血抑制率分别为42.14%和64.83%。进一步对D-b中的单体进行验证，发现单体莫诺苷能抑制红细胞溶血，2 μmol·L-1浓度的莫诺苷对红细胞的溶血抑制率为36.04%。结论六味地黄汤的活性成分糖苷提取物D-b具有抗补体活性作用，有望发展成为新型抗补体药物，需进一步进行机制研究。

关键词：中药；补体抑制；活性成分；细胞溶血；转录组

基金项目：国家自然科学基金（81803431）

通讯作者：肖智勇，E-mail：zy-xiao@163.com；周文霞，E-mail：zhouwx@bmi.ac.cn

钩藤碱对阿尔茨海默病模型斑马鱼p38 MAPK/N-cadherin的影响

刘凯菲，吴世敏，李讯怡，黄 瑶，关国凯，陈晓龙，陈毅飞

（桂林医学院药学院，广西 桂林 541199）

摘要：目的探讨钩藤碱对三氯化铝（AlCl3）诱导的阿尔茨海默病（AD）模型斑马鱼的影响。方法将斑马鱼放于T迷宫中训练7 d，选取训练成功的60条斑马鱼，分为6组，分别为：空白组、模型组、阳性组、低剂量组、中剂量组、高剂量组，每组10条。将斑马鱼适应性喂养2 d，第3天开始训练，并用记录其运动轨迹；除空白组外，将其余5组进行造模，并记录其运动轨迹；造模后，给药组给予钩藤碱给药，阳性组给予石杉碱甲给药，记录其运动轨迹，使用行为学记录分析软件Smart3.0分析其记录好的运动轨迹。提取给药后的斑马鱼脑组织，用Western印迹法检测斑马鱼脑组织P38、p-P38蛋白和N-cadherin蛋白的表达水平。结果对比分析6组斑马鱼在绿色短臂区域所待时间的数据：空白组与模型组有显著性差异（P＜0.05），模型组与阳性组、钩藤碱给药低剂量组、中剂量组、高剂量组有显著性差异（P＜0.05）。对比分析6组斑马鱼在绿色臂区域的时间所占百分比的数据：空白组与模型组有显著性差异（P＜0.05），模型组与阳性组、钩藤碱给药组低剂量组、中剂量组、高剂量组均有显著性差异（P＜0.05）。使用Western印迹法检测蛋白，相比于模型组，发现钩藤碱给药组N-cadherin蛋白表达水平上调（P＜0.05）；相比于模型组，发现空白组、阳性组、钩藤碱给药组p-P38蛋白表达水平下调（P＜0.05）。结论钩藤碱可以通过下调p-P38/P38的比值，上调N-cadherin，进而改善AD模型斑马鱼的学习记忆能力。

关键词：钩藤碱；阿尔茨海默病；p38 MAPK；N-cadherin

基金项目：广西壮族自治区自然科学基金（2017GXNSFAA198255；2018GXNSFBA138028）；广西脑与认知神经科学重点实验室开放课题（GKLBCN-20180105-03）；2019大学生创新创业项目训练计划（201910601038）；第四期广西高等学校千名中青年骨干教师培育计划

通讯作者：陈毅飞，E-mail：chenyifei_pharm@glmc.edu.cn

钩藤碱对阿尔茨海默病模型斑马鱼行为的影响

刘凯菲，吴世敏，李讯怡，黄 瑶，关国凯，黄雪虹，陈毅飞

（桂林医学院药学院，广西 桂林 541199）

摘要：目的探讨钩藤碱对三氯化铝（AlCl3）诱导的阿尔茨海默病（AD）模型斑马鱼行为的影响。方法对训练前的斑马鱼进行录像，便于判断斑马鱼是否训练成功。而后对斑马鱼进行训练，训练7 d，将训练成功的60条斑马鱼随机分为6组：空白组、模型组、阳性组、钩藤碱低剂量组、中剂量组、高剂量组。对空白组进行录像，而模型组、阳性组、低剂量组、中剂量组、高剂量组给予AlCl3染毒造模，结束后对模型组进行录像，其余各组均给予药物干预。阳性组与钩藤碱给药组均给药6 d，最后对所有给药组进行录像。全部录像完毕，用行为学记录分析软件Smart3.0进行行为学分析，通过分析数据得出结论。结果用6组斑马鱼在红色短臂区域所待时间的数据进行对比分析：空白组与模型组有显著性差异（P＜0.05），模型组与阳性组、钩藤碱中剂量组、高剂量组有显著性差异（P＜0.05）。用6组斑马鱼在红色短臂区域的时间所占百分比的数据进行对比分析：空白组与模型组有显著性差异（P＜0.05），模型组与阳性组、钩藤碱给药组中剂量组、高剂量组均有显著性差异（P＜0.05）。对6组斑马鱼在红色短臂区域游动距离的数据进行对比分析：空白组与模型组无显著性差异，而模型组只与高剂量组有显著性差异（P＜0.05）。但6组斑马鱼在红色短臂区域游动距离所占百分比这组数据对比分析：空白组与模型组有显著性差异（P＜0.05），模型组与阳性组、钩藤碱给药组中剂量组、高剂量组均有显著性差异（P＜0.05）。结论钩藤碱具有改善AD模型斑马鱼行为的作用。

关键词：钩藤碱；阿尔茨海默病；斑马鱼；行为学分析

基金项目：广西壮族自治区自然科学基金（2017GXNSFAA198255；2018GXNSFBA138028）；广西脑与认知神经科学重点实验室开放课题（GKLBCN-20180105-03）；2019大学生创新创业项目训练计划（201910601038）；第四期广西高等学校千名中青年骨干教师培育计划

通讯作者：陈毅飞，E-mail：chenyifei_pharm@glmc.edu.cn

复方利血平氨苯蝶啶片降压作用机制的网络药理学分析及验证研究

刘 珊1，2，张 雯1，王守宝1，杜冠华1，2

（1.中国医学科学院北京协和医学院药物研究所，北京市药物靶点研究与新药筛选重点实验室，北京 100050；2.广东药科大学，广东 广州 510006）

摘要：目的复方利血平氨苯蝶啶片是用于临床治疗高血压病的复方降压药，由我国学者研发，在临床应用多年，但其降压作用机制仍有待阐释。本文通过网络药理学分析其降压作用机制并在细胞水平进行初步研究验证。方法本文利用Swiss Target Prediction数据库获得复方利血平氨苯蝶啶片中利血平、氨苯蝶啶、氢氯噻嗪和硫酸双肼屈嗪4种化学成分的作用靶点；通过TTD、OMIM数据库查找并获取高血压病的相关蛋白靶标；利用蛋白互作网络分析数据库构建复方利血平氨苯蝶啶片-高血压病网络模型；通过Metascape数据库对潜在靶点进行GO富集分析和通路富集分析；采用人脐静脉内皮细胞和血管平滑肌细胞分别经血管紧张素Ⅱ1 μmol·L-1诱导处理，低中高浓度复方利血平氨苯蝶啶片原料（0.01，0.1和1 μmol·L-1）进行干预，Western印迹法检测人脐静脉内皮细胞中PI3K/Akt/eNOS通路和血管平滑肌细胞中cGMP/PKG通路的变化。结果网络药理学结果表明，复方利血平氨苯蝶啶片可能通过参与管径调节、血管张力调节、全身动脉血压的负向调节、肾上腺素受体激活、蛋白激酶活性等实现降压，降压过程中可能涉及cGMP/PKG信号通路、血管平滑肌收缩、调控神经活动的配体-受体相互作用、心肌细胞肾上腺素信号传导、钙信号通路等信号通路。细胞水平的研究结果证明，复方利血平氨苯蝶啶片处理后，可以升高人脐静脉内皮细胞中p-PI3K、p-Akt、p-eNOS以及血管平滑肌细胞中eNOS、p-VASP、PKG蛋白水平。结论复方利血平氨苯蝶啶片有效地控制血压，预防和减少心血管事件是通过多靶点、多通路发挥降压作用，进一步研究证实，激活PI3K/Akt/eNOS通路诱导内皮依赖的NO/cGMP/PKG信号，从而舒张血管可能是其重要作用机制之一。

关键词：高血压；复方利血平氨苯蝶啶片；网络药理学

通讯作者：王守宝，E-mail：shoubaowang@imm.ac.cn；杜冠华，E-mail：dugh@163.comimm.ac.cn

感冒清热颗粒对风寒表证小鼠的保护作用

刘珊宏，夏玉英，卢子滨，曹惠慧，刘俊珊，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的感冒清热颗粒具有疏风散热、清热解表的功效，临床上常用于治疗风寒感冒。本研究观察感冒清热颗粒对风寒表证小鼠模型的影响，阐明其抗风寒感冒的作用机制，为临床应用提供实验依据。方法BALB/c小鼠70只，随机分为正常对照组、风寒模型组、感冒清热颗粒3.6，7.2和14.4 g·kg-1·d-1组和阳性药物组（风寒感冒颗粒3.6 g·kg-1·d-1组和感康0.15片·kg-1·d-1组），每组10只。除正常对照组，其余组根据《中医实验动物学》采用风扇加寒冷体表降温吹风方法造模7 d，复制风寒犯表的太阳病动物模型。正常组不做任何处理饲养7 d。从开始造模第4天，药物组ig给药3 d，正常组和模型组ig给予等量生理盐水。期间记录各组动物的体重，第8天各组小鼠取血，处死后取肺、脾和胸腺，称重，计算肺、脾和胸腺的脏器系数，小鼠左肺固定后用于做病理切片HE染色，右侧肺组织匀浆检测其中肿瘤坏死因子-α（TNF-α）、白细胞介素6（IL-6）和IL-1β的水平，抗凝血用于血常规检测。结果经风寒刺激7 d，模型组小鼠体重下降（P＜0.01），感冒清热颗粒14.4 g·kg-1·d-1使其增加（P＜0.05）。模型组小鼠肺指数升高（P＜0.01），脾指数下降（P＜0.01），胸腺指数无差异，感冒清热颗粒各剂量均抑制肺指数升高（P＜0.01），对脾指数无影响。病理组织学检查显示，模型组肺间质增生，血管壁增厚，局部有出血，中性粒细胞浸润，部分区域出现代偿性空泡，感冒清热颗粒不同剂量组药物干预后缓解。模型组小鼠肺组织炎症因子TNF-α，IL-1β和IL-6表达增多（P＜0.05，P＜0.01，P＜0.01），感冒清热颗粒7.2和14.4 g·kg-1·d-1干预后炎症因子表达降低（P＜0.01）。模型组小鼠外周血中红细胞、血小板与血红蛋白数量无显著性变化，白细胞水平下降（P＜0.01），感冒清热颗粒干预后白细胞水平有所回升，但无显著性差异。在白细胞各个细胞群比例中，模型组淋巴细胞比例下降（P＜0.01），单核细胞与中性粒细胞比例升高（P＜0.01，P＜0.05），高剂量感冒清热颗粒给药后淋巴细胞、单核细胞比例向正常组回归（P＜0.05，P＜0.01）。结论感冒清热颗粒对风寒表证小鼠有保护作用，表现为增加风寒表证小鼠的体重，抑制肺指数升高，减轻肺部炎症细胞浸润与出血，其机制与降低肺组织TNF-α，IL-6和IL-1β有关。

关键词：感冒清热颗粒；风寒感冒；风寒表证模型；抗炎

通讯作者：余林中，E-mail：yulzh@smu.edu.com

7种单味中药及其组方体外抑菌效果及复方溶液脱色方法研究

刘小凤，侯雯倩，刘东玲，海 洋，苏 敬，董劲曲，徐 兰

（甘肃中医药大学药学院，甘肃 兰州 730000）

摘要：目的探究五倍子、黄芩、金银花、板蓝根、连翘、薄荷和山楂7味“药食同源”中药及其组方在大肠杆菌、金黄色葡萄球菌及白色念珠菌中的抑菌作用，筛选“药食同源”中药组方的最佳配比并初步确定该中药组方的脱色方法。方法使用水煎煮法提取各单味中药及其组方的活性成分，采用琼脂平板打孔法、纸片扩散法和平板培养法测定其抑菌圈直径、最小抑菌浓度（MIC）和最小杀菌浓度（MBC）；设计7因素2水平的正交试验，采用抑菌圈的直径作为评判指标初步确定最优抑菌配比。以最佳配方水提液为原料，用活性炭吸附法、大孔树脂脱色法、H2O2氧化脱色法及水提醇沉法4种方法对其脱色效果进行综合评价。以脱色后药液的气味及颜色变化，以3种供试菌的抑菌圈直径和MIC等作为评价指标。结果7味中药对3种供试病原菌均具有不同程度的抑菌效果，五倍子、黄芩、金银花和山楂对大肠杆菌和金黄色葡萄球菌的抑菌作用较强，其抑菌圈直径达到14.5～30.3 mm，MIC＜250 g·L-1，MBC＜500 g·L-1，且对白色念珠菌也有一定的抑制作用；板蓝根、连翘和薄荷对3种供试菌的抑菌效抑菌活性很弱甚至无抑菌效果。正交试验结果表明，7味不同配比的中药组合对大肠杆菌、金黄色葡萄球菌的联合抑菌作用具有协同作用。由连翘、山楂、板蓝根、五倍子、金银花、黄芩和薄荷的质量比为2∶1∶2∶2∶1∶2∶1组成的中药组合物对大肠杆菌和金黄色葡萄球菌的MIC为1.95和3.90 g·L-1；MBC为7.81和31.25 g·L-1，在增强抑菌效果的同时又降低了MIC和MBC；脱色结果表明75%水提醇沉法对药液的脱色效果最佳。结论多种“药食同源”中药具有抗菌作用，且其合理的配伍可以产生协同增效的效果，这将为无毒副作用和无刺激性抗菌剂的开发奠定了基础。

关键词：“药食同源”中药；抗菌；正交试验；最佳组方配比；脱色

基金项目：2020年度甘肃省高等学校产业支撑计划项目（2020C-36）；国家自然科学基金地区基金（81760789）；甘肃省教育厅“双一流”科研重点项目（GSSYLXM-05）；甘肃省教育厅优秀研究生“创新之星”项目（2021CXZX-752）

通讯作者：刘东玲，E-mail：dongling83@163.com

基于网络药理学探究三棱提取物抑制黑色素瘤的药效和分子机制

刘洋

（南京中医药大学，江苏 南京 210023）

摘要：目的基于网络药理学探究三棱提取物治疗黑素瘤的潜在分子机制，并设计实验加以验证。方法通过中药系统药理学数据库与分析平台（TCMSP）检索三棱的活性成分及对应的靶点。利用GeneCards数据库检索黑色素瘤疾病靶点，借助cytoscape软件绘制药物-疾病靶点蛋白互作（PPI）网络，并筛选出核心靶点，上传至DAVID平台进行KEGG通路富集分析，最后通过实验加以验证。结果从TCMSP数据库中获得5个三棱中活性成分，对应的靶点共70个，与疾病靶点构建PPI网络并进一步筛选得到86个核心靶点组成的关键网络，KEGG信号通路富集分析结果表明，三棱治疗黑色素瘤主要涉及糖代谢、碳代谢、氨基酸代谢等与能量代谢密切相关的通路，实验结果表明，三棱提取物降低了黑色素瘤细胞的胞外酸化率及糖酵解代谢产物的生成。结论三棱提取物防治黑色素瘤具有多成分、多靶点、多通路的特点，其中对肿瘤细胞代谢的影响主要是通过干预糖酵解途径来抑制黑色素瘤的生长的。

关键词：网络药理学；三棱提取物；黑素瘤

通讯作者：刘 洋，E-mail：20190372@njucm.edu.cn

拟人参皂苷-F11对大鼠血栓栓塞性脑卒中的防治作用及血栓性炎症相关调控机制

刘月阳，高永峰，杨静玉，吴春福

（沈阳药科大学生命科学与生物制药学院，辽宁 沈阳 110016）

摘要：目的对缺血性脑卒中发病机制的深入研究发现，血小板参与的血栓性炎症在缺血-再灌注损伤中具有重要作用。拟人参皂苷-F11（PF11）是西洋参茎和叶中的一种三萜皂苷，本课题组前期研究发现其能够显著抑制大鼠永久性和短暂性脑缺血损伤。然而，PF11能否通过调节血栓性炎症而改善栓塞性脑缺血损伤尚未见报道。方法采用大鼠血栓大脑中动脉阻塞（eMCAO）模型，结合神经行为学、Western印迹、免疫荧光等检测技术，考察PF11对eMCAO所致大鼠脑缺血损伤的改善作用及血栓性炎症相关的调控机制。结果每天单次尾静脉注射（iv）给予PF11（4，12和36 mg·kg-1），连续3 d，能够显著减小eMCAO大鼠的脑梗死体积、减轻脑水肿、改善神经功能障碍、抑制缺血侧皮质神经元的缺失以及改善缺血侧局部脑血流量。PF11（4，12和36 mg·kg-1，iv）可显著降低eMCAO大鼠缺血侧脑内血栓形成数量，减少缺血侧脑内纤维蛋白和血小板在微血管中的沉积，显著抑制大鼠缺血侧脑内炎症细胞的浸润和活化以及炎症介质的生成。进一步研究显示，PF11能够抑制eMCAO大鼠体内血管性血友病因子-糖蛋白Ib（vWF-GPIb）轴和接触-激酶通路的激活，表现为显著抑制血小板受体GPIbα和GPVI蛋白水平的上调，下调缺血侧脑内的vWF蛋白的表达，降低缺血侧皮质内FXIIa、前激肽释放酶水平和激肽释放酶生成，抑制激肽原释放缓激肽以及缓激肽1受体的上调。结论PF11可能通过抑制血栓性炎症发挥对eMCAO所致脑损伤的改善作用，其机制可能与抑制vWF-GPIb轴和接触-激酶通路的激活以及抑制血小板的黏附功能有关。

关键词：拟人参皂苷F-11；血栓栓塞性脑卒中；血栓性炎症

通讯作者：吴春福，E-mail：wucf@syphu.edu.cn；杨静玉，E-mail：yangjingyu2006@gmail.com

灵芝酸通过髓样分化蛋白2调控小胶质细胞活化改善小鼠脑缺血损伤

马 昂，王淑园，全亚竹，李 敏，杨宝学

（北京大学基础医学院药理学系，北京 100191）

摘要：目的灵芝酸是灵芝的主要生物活性成分之一，具有免疫调节及抗炎的作用。本研究旨在观察灵芝酸对小鼠脑缺血损伤是否具有保护作用及可能的作用机制。方法通过线栓法缺血60 min再灌注制备小鼠脑缺血再灌注损伤模型。小鼠随机分为假手术组、假手术给药组、模型组和灵芝酸治疗组。灵芝酸（1.25，5和20 mg·kg-1）于再灌同时ip给药。体外实验应用经典的脂多糖（LPS）诱导小胶质细胞神经炎症损伤模型。结果灵芝酸可呈剂量依赖性改善小鼠脑缺血后神经功能缺失、减小脑梗死体积、减轻脑水肿程度。与模型组相比，灵芝酸治疗组可显著抑制脑缺后神经元的凋亡，抑制小胶质细胞过度激活，减少损伤侧脑组织中炎症相关蛋白iNOS，COX-2和TNF-α的表达，NF-κB及MAPK信号通路的蛋白（ERK，JNK和P38）磷酸化水平也显著降低。体外实验结果表明，灵芝酸同样可呈剂量依赖性地减少LPS诱导后小胶质细胞炎症相关蛋白iNOS，COX-2和TNF-α的表达水平，并抑制NF-κB及MAPK信号通路。我们利用超高效液相色谱-串联质谱检测灵芝酸的组成，并将含量较高的单体进行提取和纯化，通过ELSIA法筛选有效的抗炎单体，应用表面等离子共振技术、蛋白芯片技术、分子对接技术以及免疫共沉淀等方法对筛选出的灵芝酸可能的作用靶点进行检测和验证。结果表明，灵芝酸混合物主要由19种灵芝酸单体组成，其中含量较高的8种单体均可抑制小胶质细胞炎症因子的分泌，并且髓样分化蛋白2（MD2）可能为灵芝酸抑制小胶质细胞炎症反应的关键靶点，灵芝酸单体可通过与MD2结合，抑制其与TLR4形成复合物，进而影响下游的NF-κB及MAPK信号通路。在8种灵芝酸单体中灵芝酸K的抗炎活性最为突出，且与MD2亲和力也最强。MD2基因敲除及MD2基因敲除后给予灵芝酸对小胶质细胞炎症反应及脑缺血再灌注损伤的实验结果表明，脑缺血再灌注损伤后，损伤侧脑组织中MD2的表达明显升高，MD2基因敲除后显著抑制脑缺血再灌注后小胶质细胞的过度激活。并且MD2基因敲除可抑制LPS引起的小胶质的炎症反应，减少脑缺血再灌注后脑梗死体积，在MD2基因敲除基础上给予灵芝酸，以上作用无显著差异。表明灵芝酸发挥脑缺血再灌注损伤保护作用依赖于MD2。结论灵芝酸具有抗脑缺血损伤的作用，其机制主要是通过与MD2结合，抑制MD2与TLR4形成复合物，下调MAPK及NF-κB信号通路，进而抑制小胶质细胞过度激活，减少炎症因子的表达。

关键词：灵芝酸；髓样分化蛋白2；小胶质细胞；脑缺血损伤

通讯作者：杨宝学，E-mail：baoxue@bjmu.edu.cn

白头翁皂苷B4抑制香烟烟雾诱导的慢性阻塞性肺病及癌前病变的作用

马惠苗1，2，3，陈兰英1，2，3，周子也1，2，3

〔1.中药固体制剂制造技术国家工程研究中心，江西南昌330006；2.江西中医药大学，江西南昌 330004；3.中药药效（扶正祛邪抗肿瘤）评价重点研究室，江西南昌 330006〕

摘要：目的研究白头翁皂苷B4对香烟烟雾（CS）诱导的慢性阻塞性肺病（COPD）及癌前病变的影响并探讨作用机制。方法80只C57BL/6小鼠，雌雄各半，通过连续CS诱导法建立COPD小鼠模型。在造模第9周，按照体重将小鼠随机分为正常组、CS诱导组、白头翁皂苷B4 1.5，3和6 mg·kg-1组，阳性对照药地塞米松1.5 mg·kg-1组。熏烟前1 h给药，持续4周。造模及给药结束后，采用肺功能仪测定小鼠肺功能参数的变化；高通量蛋白芯片技术测定支气管肺泡灌洗液中炎症因子的含量；HE染色观察肺组织病理变化及上皮细胞增生情况；PCR法测定小鼠肺组织中转化生长因子β（TGF-β）、肿瘤坏死因子α（TNF-α）、白细胞介素1β（IL-1β）基因表达水平以及基质金属蛋白酶2（MMP2），MMP9，MMP12和MMP抑制剂（TIMP1）的表达；采用试剂盒测定小鼠血浆中髓过氧化物酶（MPO）和谷胱甘肽过氧化物酶（GSH-Px）活性以及丙二醛（MDA）含量；免疫组织化学法测定肺组织中抑癌基因（p53）、增殖细胞核抗原（ki67）以及核内不均一核糖核蛋白（hnRNP）的表达；Western印迹法检测小鼠肺组织中DNA甲基转移酶（Dnmt1）、脆性组氨酸三联体（FHIT）、抑癌基因CDKN2A/P16蛋白表达水平。结果白头翁皂苷B4能够显著降低吸气时间（Ti）、肺阻力（LR）（P＜0.05），升高最大呼气流速（PEF）（P＜0.05）：白头翁皂苷B4可缓解炎症细胞浸润情况，并能够降低支气管肺泡灌洗液中炎症因子IL-2，IL-5，IL-18和IL-10含量（P＜0.01），显著下调小鼠肺组织中炎症因子TGF-β，TNF-α和IL-1β基因表达水平（P＜0.01），显著降低MMP2和MMP12基因表达，升高MMP9、TIMP1基因表达水平（P＜0.01）；白头翁皂苷B4给药组能够不同程度降低MDA含量及MPO活性，同时能够显著提升GSH-Px活性（P＜0.05，P＜0.01）；免疫组织化学实验结果表明，白头翁皂苷B4 3和6 mg·kg-1组均能够显著上调P53蛋白表达（P＜0.01）；白头翁皂苷B4对Dnmt1和Cdkn2a/P16蛋白表达无影响，可显著下调FHIT蛋白表达水平（P＜0.01）。结论白头翁皂苷B4对COPD小鼠肺功能具有一定程度改善作用，能够通过平衡小鼠肺组织中蛋白酶与抗蛋白酶表达，降低氧化应激水平，改善肺组织氧化应激进而起到保护作用，并有抑制癌前病变的作用趋势。

关键词：白头翁皂苷B4；香烟烟雾提取物；慢性阻塞性肺病；癌前病变

基金项目：江西省重点研发计划（20181ACG70014）；国家自然科学基金（82060732）；江西省中医药重点研究室建设项目

通讯作者：陈兰英，E-mail：clyxy2513@163.com

沉香提取物保护胆汁酸诱导胃黏膜损伤的药理机制

马家乐，王鑫玉，靳凤玉，赵一慕，史景景，郑 姣，李 军

（北京中医药大学中药学院中药现代研究中心，北京 100029）

摘要：目的胃黏膜上皮细胞损伤是胆汁反流性胃炎最重要的病理特征之一。沉香在我国传统用药中用于治疗胃病已有数千年的历史。然而胆汁酸诱导胃黏膜上皮细胞损伤的病理机制以及中药沉香改善胆汁反流性胃炎的作用靶点尚未完全阐明。本研究探讨牛磺胆酸（TCA）通过调节内质网应激途径促进细胞凋亡的作用，并探讨了沉香提取物（CPE）对胃黏膜上皮细胞的保护作用和作用机制。方法利用TCA（0.5，1.0，1.5，2.0 μmol·L-1）处理人胃黏膜上皮细胞GES-1细胞24 h后，利用CCK-8试剂盒检测GES-1细胞存活率，确认TCA诱导细胞损伤的IC50为1.467 μmol·L-1。选择1.5 μmol·L-1的TCA诱导GES-1细胞损伤，同时采用CPE（1，5，10 mg·L-1）处理细胞后，检测CPE对细胞存活率的改善作用。利用Hoechst33342染色和Annexin V-FITC/PI双染流式细胞术检测TCA对细胞凋亡的激活以及CPE的改善作用。通过Western印迹法检测胱天蛋白酶7、活化胱天蛋白酶7、PARP、活化PARP等凋亡蛋白表达情况。进一步通过Western印迹法检测内质网应激相关蛋白Bip，PERK，eIF2α，CHOP的表达，探讨胆汁酸处理和CPE对内质网应激的影响。体内实验采用胆汁反流性胃炎小鼠检测体重变化、血浆炎症因子IL-6，IL-1β，TNF-α水平，采用HE染色、TUNEL染色、免疫组化进一步验证CPE对胆汁反流性胃炎的治疗作用。结果TCA剂量依赖性地降低GES-1细胞的存活率（IC50=1.467 μmol·L-1），CPE 5和10 mg·L-1显著增加TCA损伤细胞的存活率（P＜0.05）。Hoechst33342染色和流式细胞术分析结果显示，TCA处理后细胞凋亡率显著提高，CPE给药后剂量依赖性抑制了TCA诱导的细胞凋亡（P＜0.05）。Western印迹法结果显示，TCA处理GES-1细胞后，通过增加胱天蛋白酶7和PARP的裂解激活了GES-1细胞的凋亡。进一步的实验表明，TCA可上调内质网应激继而诱导细胞凋亡，CPE抑制内质网应激相关蛋白PERK和eIF2α的磷酸化，降低CHOP的表达水平，从而发挥其抗凋亡作用。体内实验胆汁反流性胃炎小鼠血浆炎症因子水平以及胃黏膜上皮细胞凋亡和CHOP蛋白水平也明显增加，CPE给药后明显降低。CPE通过抑制PERK/eIF2α/CHOP信号通路，有效抑制TCA激活的GES-1细胞凋亡，并且CPE对胃黏膜的抗凋亡作用也在体内得到验证。结论CPE通过调控PERK/eIF2α/CHOP信号通路介导的内质网应激抑制TCA诱导的胃黏膜上皮细胞凋亡，为胆汁反流性胃炎的治疗提供了潜在的药物。

关键词：沉香；胆汁反流；胆汁酸；胃黏膜损伤；内质网应激

基金项目：国家重点研发项目（2018YFC1706400）；国家自然科学基金（82003912）

通讯作者：李 军，E-mail：drlj666@163.com；郑 姣，E-mail：zj98v2@163.com

北沙参多糖对免疫系统调节作用研究进展

邱晓月1，2，景永帅3，郑玉光1，2，吴兰芳1，2

（1.河北中医学院药学院，河北 石家庄 050200；2.河北省中药炮制技术创新中心，河北 石家庄 050091；3.河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：北沙参（Radix Glehniae）来源于伞形科植物珊瑚菜（Glehnia littoralis Fr.Schmidt ex Miq.）的根，是临床上常用中药。本品性甘、微苦、性微寒，归肺、胃经。有益胃生津、养阴清肺之功效，主要用于肺热燥咳、劳嗽痰血、胃阴不足、热病津伤、咽干口渴等症。研究表明，北沙参所含的主要成分有香豆素、木脂素、聚炔类、多糖类等。其中多糖类成分为其活性成分之一，多糖类成分含量最高，具有抗氧化、降血糖和调节机体免疫等功能。北沙参多糖主要通过以下3方面发挥对免疫系统的调节作用：①对免疫系统和器官的作用。北沙参多糖对可使甲亢型阴虚小鼠体质量明显增加，显著增强甲亢型阴虚小鼠脾NK细胞杀伤率和T淋巴细胞转化功能，提高血清IgM和IgG含量，增强非特异性免疫和特异性免疫功能；采用小鼠碳粒廓清实验得到北沙参多糖有增强小鼠吞噬碳粒的K值以及α指数的作用；北沙参多糖可改善环磷酰胺引起的小鼠脾指数下降，进而实现对小鼠免疫功能的调控。②体外对免疫细胞的作用。北沙参多糖对脾淋巴细胞的增殖具有促进作用，且呈一定的剂量依赖性。另外北沙参多糖可增强巨噬细胞的吞噬能力，并促进分泌IL-6和TNF-α的能力，促进一氧化氮合酶（iNOS）mRNA表达水平，iNOS的提高进而诱导细胞产生NO。③对免疫分子的作用。北沙参多糖对荷瘤小鼠脾淋巴细胞增殖的促进作用和对TNF-α和IFN-γ分泌的增强作用与其能激活TLR4有关；其机制可能是多糖通过与巨噬细胞表面的TLR4结合，再经P38/MAPK信号转导途径激活下游NF-κB，从而促进TNF-α的产生。另外北沙参多糖可与巨噬细胞表面CR3结合后，可激活PI3K和MAPK信号通路，促进炎症因子的生成，发挥其免疫调控作用。综上，北沙参多糖对机体免疫系统的调控作用具有多靶点和普遍存在于特异性及非特异性免疫激活过程的各个环节的特点，免疫调控机制主要是通过靶向作用于免疫细胞表面Toll样受体、补体受体等受体分子，激活MAPK和NF-κB等下游信号通路，促进NO，TNF-α，IL-6及IFN-γ等细胞因子的分泌表达，进而调控免疫系统发挥免疫调控活性。

关键词：北沙参；多糖；免疫

基金项目：河北省省级科技计划（H2021423057）

通讯作者：吴兰芳，E-mail：wulanfang757@163.com

银杏蜜环口服溶液对大鼠心肌梗死后抑郁的干预作用

任建勋1，徐 立1，王益民2，王 勇2，顾国强2，赫少清2，刘 赛1，刘建勋1

（1.中国中医科学院西苑医院基础医学研究所，北京 100091；2.西安步长中医心脑病医院，陕西 西安 710082）

摘要：目的观察银杏蜜环口服溶液（简称“银蜜”）对冠状动脉左前降支结扎以及温和的不可预见性刺激诱导大鼠心肌梗死后抑郁的干预作用。方法大鼠按体重随机分为空白对照组，心肌梗死组，心肌梗死后抑郁模型组，心肌梗死后抑郁阳性药物组（氟哌噻吨美利曲辛片，0.44 mg·kg-1），银蜜3和6 mL·kg-1组，每组10只。采用冠状动脉左前降支结扎建立心肌梗死模型后ig给予不同药物，同时采用温和不可预见性刺激建立大鼠抑郁模型，持续28 d。实验4周后，分别测定各组大鼠体重、进食量、强迫性游泳、糖水偏好程度和旷场实验评价大鼠与抑郁相关的行为学变化；采用心脏超声评价大鼠心脏结构和功能改变；采用心肌组织染色评价心室重构程度；并检测血清中皮质酮、氧自由基（OFR）和总抗氧化能力（T-AOC）水平变化。结果与空白对照组比较，模型组大鼠体重4周时明显下降；同时蔗糖水消耗和摄食量明显下降（P＜0.01）；强迫性游泳不动时间明显增加（P＜0.01）；自主活动中大鼠总路程和平均速度都有明显下降（P＜0.01）；同时模型组大鼠LVAWs，LVAWd，LVPWs和LVPWd明显降低，而LVIDs，LVIDd，LVEVs和LVEVd明显升高（P＜0.05或P＜0.01）；此外模型组大鼠SV，EF，FS和CO也明显下降（P＜0.05或P＜0.01）。室腔面积占心肌总面积的百分比明显升高。模型组大鼠血清皮质酮和OFR明显升高，而血清T-AOC则明显下降（P＜0.01）。与模型组比较，银蜜6 mL·kg-1组大鼠体重在4周时明显升高；同时银蜜6 mL·kg-1组和心肌梗死组大鼠蔗糖水消耗和摄食量有一定程度地增加，阳性药大鼠蔗糖水消耗也明显增加（P＜0.05）；同时银蜜6 mL·kg-1组、阳性药组以及心肌梗死组大鼠强迫性游泳不动时间明显减少（P＜0.05）；而阳性药组大鼠活动的总路程和平均速度，以及心肌梗死组大鼠活动的总路程都明显升高（P＜0.05）。与模型组相比，银蜜6 mL·kg-1组、阳性药组和心肌梗死组大鼠LVPWs升高，同时银蜜6 mL·kg-1组大鼠LVAWs和LVPWd以及SV，EF，FS和CO也明显增加（P＜0.05或P＜0.01），而左心室腔面积占心肌总面积的百分比明显下降（P＜0.05）。与模型组大鼠相比，银蜜3和6 mL·kg-1组、阳性药组和心肌梗死组大鼠血清皮质酮含量下降（P＜0.05），银蜜3和6 mL·kg-1组大鼠血清T-AOC明显上升，而血清OFR含量都有明显下降（P＜0.05或P＜0.01）。结论采用冠状动脉左前降支结扎以及温和的不可预见性刺激的方法能够很好的建立大鼠心肌梗死后抑郁模型。银杏蜜环口服溶液能够改善心肌缺血后心室重构及心脏功能；并改善心肌梗死后抑郁大鼠的生理和行为学的改变。与银杏蜜环口服溶液减轻心肌梗死后抑郁过程中出现的氧化应激反应有一定的关联。

关键词：银杏蜜环口服溶液；心肌梗死；抑郁

通讯作者：刘建勋，E-mail：liujx0324@sina.com

川续断皂苷B抑制线粒体E3泛素连接酶1减轻缺血性中风大鼠脑损伤

任凯迪1，彭紫梅1，罗秀菊2，彭 军1

（1.中南大学湘雅药学院药理学系，心血管研究湖南省重点实验室，湖南 长沙 410078；2.中南大学湘雅三医院检验科，湖南 长沙 410013）

摘要：目的探讨川续断皂苷B对缺血性脑卒中的作用及机制。方法SD大鼠和大鼠肾上腺嗜铬细胞瘤PC12细胞分别给予左脑缺血2 h、再灌注24 h和低氧8 h、复氧24 h的处理，在动物水平和离体水平建立缺血性脑卒中模型并给予不同剂量的川续断皂苷B。通过神经功能学评分、TTC染色、TUNEL染色检测大鼠脑损伤以及运用MTS法、LDH细胞毒性释放实验、流式细胞术等手段检测PC12细胞损伤，并运用Western印迹法对相关蛋白水平进行检测。运用MOE分子对接软件对川续断皂苷B和线粒体E3泛素连接酶1（Mul1）进行分子相互作用预测。结果与假手术组相比，模型组大鼠发生了严重脑损伤（神经功能学评分升高、脑梗死灶出现、TUNEL阳性细胞增多，胱天蛋白酶3活性增强、ATP产量减少），同时Mul1和动力相关蛋白1（Drp1）水平升高，mitofusin2（Mfn2）蛋白水平降低。川续断皂苷B能够减弱脑损伤并逆转Mul1，Drp1和Mfn2的蛋白水平改变。离体水平结果一致，与对照组相比，低氧处理的PC12细胞出现了细胞损伤（细胞PI阳性率从5%左右增加至35%左右、胱天蛋白酶3活性增强、LDH释放率增加）和线粒体功能下降（线粒体膜电位ΔΨm和ATP产量下降，ROS产量和线粒体分裂增加），同时Mul1和Drp1表达上调，Mfn2蛋白水平降低，以上现象能被川续断皂苷B抑制。MOE分子对接结果显示，川续断皂苷B可能和Mul1 294位Asp、303位和320位Val和318位Gly存在相互作用。结论川续断皂苷B能够通过抑制Mul1改善线粒体功能保护大鼠脑组织免于缺血性损伤。

关键词：川续断皂苷B；缺血性脑卒中；线粒体E3泛素连接酶1；动力相关蛋白1；mitofusin2

通讯作者：彭 军，E-mail：jungpeng@csu.edu.cn；罗秀菊，E-mail：xjluo22@csu.edu.cn

基于蛋白质组学探究鸦胆子油下调Ras/MAPK信号通路的抗肿瘤机制

苏 敬，侯雯倩，刘小凤，董劲曲，徐 兰，刘东玲，海 洋

（甘肃中医药大学，甘肃 兰州 730000）

摘要：目的基于蛋白质组学以秀丽隐杆线虫为模式生物探究鸦胆子油的急毒性及抗肿瘤机制。方法提取、制备鸦胆子油乳，稀释成不同浓度后加入96孔板，置于恒温摇床中分别培养1 d观察线虫的死亡比率；培养5 d后观察野生型线虫的比率，分析鸦胆子油乳的急毒性与抗肿瘤活性。提取线虫总蛋白，运用串联质谱标签定量蛋白质组学研究药物干预前后的差异蛋白情况。结果鸦胆子油的毒性较小，浓度为25 mg·L-1时，线虫的死亡率为46%；与对照组相比，给药组均对Ras基因突变的线虫多阴门表型有不同程度的抑制作用，且随着浓度的增大，其对秀丽隐杆线虫的多阴门表型的抑制作用增强；蛋白质组学数据分析筛选出127个差异蛋白，其中上调蛋白97个，下调蛋白30个；GO功能富集显示，主要与DNA复制、DNA链延伸、染色质沉默的负调控、单链DNA解旋酶活性、氧化还原酶活性等功能有关。KEGG富集显示主要与TGF-β信号通路、Wnt信号通路、Hedgehog信号通路、MAPK信号通路等多条信号通路相关。结论鸦胆子油毒性较小，且能够明显下调Ras/MAPK通路，具有较好的抗肿瘤活性。

关键词：鸦胆子油；秀丽隐杆线虫；Ras/MAPK；蛋白质组学

基金项目：国家自然科学基金（81760789；82074419）；甘肃中医药大学科创基金重点项目

通讯作者：刘东玲，E-mail：dongling83@163.com；海 洋，E-mail：cathyhaiyang@163.com

苦参碱抗肿瘤作用及机制研究进展

苏紫藤1，景永帅1，张丹参1，郑玉光2，3，吴兰芳2，3

（1.河北科技大学化学与制药工程学院，河北 石家庄 0500182；2.河北中医学院药学院，河北 石家庄 050200；3.河北省中药炮制技术创新中心，河北 石家庄 050091）

摘要：肿瘤是指机体在各种致癌因子的作用下，局部组织细胞遗传基因改变，细胞异常增生所形成的肿块状新生物。目前治疗肿瘤的方法包括放疗、化疗以及外科手术等，但在放疗、化疗治疗过程中应用的大部分抗肿瘤药物使患者遭受巨大痛苦，并对人体产生剧烈的毒副作用。苦参碱（matrine）为豆科植物苦参、苦豆子、广豆根等中草药中的主要活性成分，具有抗炎、抗病毒、抗心律失常等多种药理作用。近年来，国内外大量研究报道，苦参碱在抗肿瘤方面也具有明显功效，新近研究的苦参抗肿瘤作用机制主要包括以下方面：①抑制肿瘤细胞增殖。研究发现，在慢性炎症部位的肿瘤发生率较非炎症部位明显增高，并且在炎症组织中过表达的细胞因子已被证实可诱导肿瘤的发生发展，炎症细胞及相关因子在肿瘤的发展过程中，白细胞介素（IL）-6是其共同作用的靶分子，IL-6可激活信号传导及转录激活蛋白（STAT3），活化的STAT3可触发靶基因的转录和表达，调控肿瘤细胞增殖及凋亡，IL-6/STAT3信号通路在部分肿瘤组织中可促进其增殖、抑制其凋亡，苦参碱可以抑制IL-6/STAT3蛋白表达，降低炎症反应，达到有效抑制肿瘤细胞增殖的作用。②调控肿瘤细胞凋亡。Bax蛋白可以促进肿瘤细胞的凋亡，Bcl-2蛋白的作用与之相反，研究表明苦参碱的抗肿瘤作用可能与抑制Bcl-2蛋白、增加Bax水平有关。FAS/FASL死亡受体介导信号通路是一种新近发现的细胞凋亡途径，其可以激活凋亡相关蛋白含胱天蛋白酶8，胱天蛋白酶8可进一步激活执行死亡功能的胱天蛋白酶3，6和7等，研究表明苦参碱可以上调FAS/FASL的表达，激活胱天蛋白酶8，从而抑制肿瘤细胞生长、促进细胞凋亡，且具有浓度依赖性。③抑制肿瘤细胞转移和侵袭。Warburg效应是指癌变细胞会偏向使用糖酵解来代替正常的有氧呼吸，Warburg效应还可以促进肿瘤上皮间质转化（EMT）的发生，EMT是肿瘤细胞发生转移和侵袭的早期标志。苦参碱可以下调糖酵解关键酶己糖激酶（HK）、丙酮酸脱氢酶（PDH）、柠檬酸合酶（CS）的表达，从而抑制肿瘤细胞的转移和侵袭。研究发现肿瘤转移的促进因子例如基质金属蛋白酶9（MMP-9）、血管内皮生长因子（VEGF）与Wnt/β-连环蛋白信号通路密切相关，苦参碱通过抑制Wnt/β-连环蛋白信号通路来降低MMP-9和VEGF的分泌，进而抑制肿瘤细胞的转移和侵袭。④增加肿瘤细胞对放疗药物的敏感性。p21激活蛋白激酶6（PAK6）是一种丝氨酸/苏氨酸激酶，能够调控细胞形态维持、骨架支撑等，可促进肿瘤的发生发展，Wnt/β-连环蛋白信号通路是肿瘤细胞内经典的信号转导通路，可调控肿瘤的生长和转移，与PAK6的表达密切相关。与单独放疗相比，苦参碱联合放疗后PAK6蛋白表达下调更加显著，其机制可能是抑制PAK6表达和Wnt/β-连环蛋白信号通路，从而提高对放疗药物的敏感性，起到治疗作用。本文综述了苦参碱抗肿瘤的作用机制，以期为抗肿瘤药物的研发和临床应用提供参考。

关键词：苦参碱；抗肿瘤

基金项目：河北省省级科技计划（H2021423057）

通讯作者：吴兰芳，E-mail：wulanfang757@163.com

四氢姜黄素的神经保护作用研究进展

孙丽丛，景永帅，张丹参

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：姜黄作为姜科植物姜黄（Curcuma longa L.）的干燥根茎，其主要化学成分有姜黄素（CUR）和挥发油等，其药理活性广泛，而四氢姜黄素（THC）作为姜黄素在生物体内的主要代谢产物，近年来也颇受关注。报道发现四氢姜黄素具有神经保护、抗癌、降血糖、降血脂等药理学作用，而关于四氢姜黄素在神经保护方面的研究也越来越多。①阿尔茨海默病（AD）：Ras/ERK信号通路是细胞生存、分化、增殖、代谢和运动受细胞外信号控制的主要机制，并与细胞周期中的G1/S转换直接相关。THC可以上调Gab2和K-Ras的表达，减轻Ccnd2的下调，通过Ras/ERK信号通路减弱G1/S阻滞的发生以及抑制小胶质细胞的细胞周期；THC下调肿瘤坏死因子α的表达，上调TGF-β1表达，发挥出抑制细胞周期停滞和凋亡的联合作用。另外THC还能降低活性氧的水平进而保护海马细胞，以及抑制胱天蛋白酶被激活，发挥抗AD的作用。②脑缺血再灌注（I/R）损伤：ERK信号通路在I/R损伤中具有重要作用，其参与生长和分化，由生长因子受体介导，在某些病理条件下还被氧化应激、脑缺血和神经递质释放激活。在目前的研究中，缺血小鼠的p-ERK水平和p-ERK/ERK比率显著增加，而THC可以抑制I/R损伤诱导的ERK途径的激活，并且这种效应是剂量依赖性的，而且还使GRASP65的磷酸化剂量依赖性降低，这为THC的神经保护特性提供了进一步的证据。③创伤性脑损伤（TBI）：在脑损伤模型中，自噬与凋亡同时发生，而自噬是一个高度调节的过程，细胞成分在饥饿过程中被降解和回收，THC通过降低细胞色素c的同型半胱氨酸化来改善自噬。细胞质中受损的细胞器和功能失调的蛋白质在自噬过程中被包装成膜泡，然后被蛋白酶体处理并最终降解，THC通过激活自噬，达到神经保护的作用。④脊髓损伤：PI3K/Akt/mTOR信号转导途径是受体信号转导细胞的重要途径，可调节细胞分化和增殖，并抑制细胞凋亡。在正常生理条件下，PI3K的细胞内表达往往较低。当细胞受损时，其表达迅速增加，导致磷脂酰肌醇磷酸化生成磷脂酰肌醇3，4-二磷酸，该通路被激活，发挥抗细胞凋亡作用。在大鼠脊髓损伤模型中，THC抑制脊髓中的水积聚，并降低炎症因子。此外，THC还可抑制氧化应激和凋亡（抑制胱天蛋白酶3活性和Bax蛋白水平），有效降低基质金属蛋白酶3和13以及环氧化酶2的基因表达，促进Akt磷酸化，增强（FOX）O4蛋白表达，达到保护脊髓的效果。

关键词：四氢姜黄素；神经保护

基金项目：河北省重点研发计划项目生物医药专项（20372509D）；河北省高等学校科学技术研究项目（ZD2020117）

通讯作者：张丹参，E-mail：zhangds2011@126.com

“明之黄芪汤”的应用历史及文献分析

谭 为1，2，杜冠华3，顾政一2

（1.新疆医科大学药学院，新疆 乌鲁木齐 830011；2.新疆维吾尔自治区药物研究所，新疆 乌鲁木齐 830004；3.中国医学科学院药物研究所，北京市药物靶点研究与新药筛选重点实验室，北京 100050）

摘要：“明之黄芪汤”原名“神效黄芪汤”，源自李东垣的《兰室秘藏》，由黄芪、人参、白芍、炙甘草、陈皮、蔓荆子组成。“明之黄芪汤”在历代医籍中皆有收载，涉及医经、医论、医案、诊法、本草、方书等，按照朝代分别为元代3部、明代23部和清代17部，虽然在炮制、剂量以及主治方面略有不同，但组方均以黄芪为主药，其它药味相辅，用以治疗痹病及眼目之证。故认为方义衍变至今，后世对本方传载中的组成、用药及主治并无太大的差异，传承过程较为连续，疗效受到历代医家认可。本方是李东垣补中益气汤、益气聪明汤化裁而来，组方原则是补中益气升阳。黄芪、人参、炙甘草补气虚；白芍养血柔肝，敛阴缓急；蔓荆子祛风升阳。根据素问至真要大论：损者益之、劳者温之的治疗原则，采用“明之黄芪汤”补益中气，使元气内充，清阳得升，诸症自除。现代应用“明之黄芪汤”多为原方的加减方，主要用于治疗神经系统疾病引起的麻木，药物剂量根据主治证候的不同也有所调整。通过重用生黄芪增加补气之功，酌加当归理血，用以治疗前部缺血性视神经病变；与指迷茯苓丸合用加减，治疗慢性酒精中毒性Wernicke脑病；与七星针局部叩刺并合方加减牵正散、当归补血汤，治疗罕见病昂白氏病5例。通过梳理和分析古代医籍与现代文献，明确了“明之黄芪汤”的应用历史，处方依据以及现代临床应用。但文献考证仅为明之黄芪汤研究开发的第一步，后续将加快明之黄芪汤指纹图谱的建立以及多组分的定性与定量研究，加快对明之黄芪汤的药效筛选，为明之黄芪汤的现代研究提供理论依据。

关键词：明之黄芪汤；历代医籍；麻木；痹症；目症

通讯作者：杜冠华，E-mail：dugh@imm.ac.cn

六味地黄苷糖对急性应激致小鼠模式分离能力损伤的影响及机制

王 昊1，2，王子元1，2，黄 晏1，2，周文霞1，2

（1.军事科学院军事医学研究院毒物药物研究所，北京 100850；2.国家抗毒药物和毒理学重点实验室，北京 100850）

摘要：目的模式分离（pattern separation）是海马脑区编码相似记忆表征并区分相似记忆信息的过程，对记忆准确性有着重要作用。急性应激可导致模式分离能力受损。六味地黄苷糖（LW-AFC）可改善应激所致的认知功能损伤，但对模式分离能力的影响尚不清楚。本研究将考察LW-AFC对急性应激模型小鼠模式分离能力的影响及其可能机制。方法本研究选用7周龄C57BL/6J小鼠，应激前连续ig给予LW-AFC 7 d（对照组给与等体积生理盐水），然后连续3 d足底电击复制急性应激模型；应激后第1天开始对动物的空间和时间模式分离进行测试；行为学检测后取血清检测皮质酮和去甲肾上腺素（NE）。结果造模后第1天，与对照组相比，模型动物的时间分辨指数有降低趋势，空间分辨指数显著降低（P＜0.05）；与模型组相比，LW-AFC组小鼠的时间分辨指数有升高趋势，空间分辨指数显著升高（P＜0.05）。造模后第8天，与对照组相比，模型组动物的时间分辨指数无明显变化，空间模式分辨指数显著降低（P＜0.05）；与模型组相比，LW-AFC组小鼠的空间分辨指数有升高趋势。血清应激激素检测结果表明，与对照组相比，模型组动物血清皮质酮含量显著升高（P＜0.05），NE含量有降低趋势；与模型组相比，LW-AFC组小鼠的血清皮质酮含量显著降低（P＜0.05），NE含量有升高趋势。结论LW-AFC能够明显改善急性应激所致的空间模式分离能力损伤，调节血清中皮质酮和NE水平可能是其改善急性应激致空间模式分离能力损伤的环节之一。

关键词：六味地黄苷糖；急性应激；模式分离；皮质酮；去甲肾上腺素

通讯作者：黄 晏，E-mail：huangyan023@163.com；周文霞，E-mail：zhouwx@nic.bmi.ac.cn

CA-30调节肠道菌群改善SAMP8小鼠情绪及学习记忆能力的作用及机制

王健辉1，2，雷 曦1，2，谢宗杰1，2，程肖蕊1，2，周文霞1，2，张永祥1，2

（1.军事科学院军事医学研究院毒物药物研究所，北京 100850；2.抗毒药物与毒理学国家重点实验室，北京 100850）

摘要：目的肠道微生物的组成与功能的改变能够影响阿尔茨海默病（AD）的疾病进程，同时还影响着情绪、认知等功能，发挥着与大脑类似的功能。前期研究发现六味地黄苷糖（LW-AFC）能够改善AD动物模型认知和情绪损伤。CA-30作为LW-AFC的寡糖部位，是调节肠道微生物组成和代谢活性最常用的物质之一。因此，本研究的目的在于揭示LW-AFC中的CA-30改善SAM小鼠肠道失调的具体生物学过程。方法以6月龄雄性SAMP8小鼠为模型动物，以SAMR1小鼠为对照动物，将SAMP8小鼠与SAMR1小鼠同笼饲养6个月。采用自主活动性、筑巢实验、老化度评分、旷场实验、高架十字迷宫以及Morris水迷宫实验分别评价小鼠的自主活动性、日常活动能力、老化程度、焦虑情绪以及空间学习记忆能力，采用16S rRNA测序技术观察小鼠肠道菌群的物种组成。应用免疫组织化学和荧光染色检测小鼠肠道结构和功能的变化。最后，ig给予6月龄雄性SAM小鼠CA-30 5个月，观察CA-30对小鼠认知、情绪及肠道菌群的影响。结果SAMR1和SAMP8小鼠同笼饲养后，显著增加了SAMR1小鼠的老化度评分和焦虑样行为，显著降低了SAMR1小鼠的空间学习记忆能力，但对SAMP8小鼠无显著影响。此外，通过肠道菌群测序发现，同笼饲养后的SAMR1肠道内Rikenella丰度显著升高，Ruminococcaceae和Cyanobacteria丰度显著降低。应用过碘酸希夫反应（PAS）和免疫荧光染色发现，同笼饲养能够显著降低SAMR1小鼠的杯状细胞数量、肠绒毛长度、肠绒毛长度/肠隐窝深度比值、紧密连接蛋白Claudin 1和胞质紧密粘连蛋白ZO-1的表达量，并且提高肠隐窝深度。CA-30能够显著影响Rikenella，Ruminococcaceae和Cyanobacteria的丰度，并能够改善SAMP8小鼠的认知功能和情绪异常。结论同笼饲养后SAMR1小鼠肠道菌群的结构和功能出现异常，导致肠黏膜受损、通透性上升，进而影响其认知功能和情绪，CA-30能够通过调节Rikenella，Ruminococcaceae和Cyanobacteria的丰度改善认知功能和情绪异常。

关键词：六味地黄方；CA-30；肠道菌群；认知功能；情绪

基金项目：中国博士后科学基金面上项目（2017M613413）；国家自然基金（81473191）

通讯作者：周文霞，E-mail：zhouwx@bmi.ac.cn

生物碱调节血脂的研究进展

王凌潇，李 军，郑 姣

（北京中医药大学中药学院，北京 100029）

摘要：生物碱是广泛存在于自然界的一类含氮有机化合物，大多数具有十分复杂的环状结构，氮元素多包含在环内，具备类似碱的性质，过去又称为赝碱，具有显著的生物活性，是中草药中十分重要的有效成分之一。高脂血症属于常见的慢性疾病，是指脂肪代谢或转运异常使血浆一种或多种脂质异于正常所引起的严重威胁人类健康的代谢性疾病的统称。血脂异常临床表现为血浆甘油三酯（TG）、总胆固醇（TC）和低密度脂蛋白胆固醇（LDL-c）异常升高；高密度脂蛋白胆固醇（HDL-c）降低。现代药理学研究表明生物碱具有调节血脂的功效，通过阻止胆酸或胆固醇从肠道吸收，促进胆酸或胆固醇从粪便排出；抑制胆固醇的体内合成；促进胆固醇的转化增强细胞膜上LDL受体表达，增加LDL-c清除，加速脂蛋白分解；激活脂蛋白代谢酶类，促进TG的水解；阻止其他脂质的体内合成，或促进其他脂质的代谢等途径达到降血脂的效果。目前国内外研究现状显示具有调节血脂异常的生物碱主要有异喹啉类生物碱、酰胺类生物碱、季胺碱以及生物碱提取部位等，本文根据作用机制对近年来生物碱调节血脂功效的文献进行综述研究，进而为生物碱治疗高脂血症的进一步研究提供理论参考。

关键词：生物碱；血脂异常

基金项目：国家自然科学基金（82074050）

通讯作者：郑 姣，E-mail：zj98v2@163.com

隧道纳米管：神经退行性疾病的新药理学靶点

王晓彤，苑玉和，陈乃宏

（天然药物活性物质与功能国家重点实验室，中国医学科学院药物研究所，北京 100050）

摘要：多样的细胞间通讯方式是维持体内平衡和传播疾病状态的重要环节。作为细胞间桥的一员，2004年发现的隧道纳米管（TNT）因其具有促进复杂细胞间通讯的作用，而被认为是潜在的药理学靶点，与多种神经退行性疾病发病机制密切相关。神经退行性疾病中常见的神经毒性蛋白聚集体包括痒病朊病毒蛋白（PrPSc）、突变tau蛋白、β-淀粉样蛋白（Aβ）、α-突触核蛋白（α-syn）以及突变亨廷顿（mHTT）蛋白。这些蛋白聚集体可以通过复杂的机制促进TNT形成，随之TNT在神经变性过程中介导这些聚集体在细胞间的传递。本综述详细描述了TNT的发现过程、骨架组成、形成机制和理化特性。同时，也详细阐述了TNT在促进多种神经退行性疾病病理发展中的关键作用，为神经退行性疾病的药物治疗提供了新的见解。

关键词：隧道纳米管；神经毒性；蛋白聚集体；细胞间通讯；神经退行性疾病

通讯作者：苑玉和，E-mail：yuanyuhe@imm.ac.cn；陈乃宏，E-mail：chennh@imm.ac.cn

六君子汤治疗神经性厌食研究进展

吴熙培，聂 克

（广东药科大学中药学院，广东 广州 510006）

摘要：神经性厌食（AN）是以强烈拒绝摄食、体重明显减轻以致危及生命为特征，并伴有肥胖恐惧及体像障碍的身心疾病。文献报道，AN的发病机制主要包括遗传、神经内分泌失衡、脑奖赏系统病变及社会心理等方面。目前的西医临床研究中，食欲刺激、药物干预和心理治疗（如家庭干预）是治疗AN的最常用方法。新近研究发现，AN不但难以根治，而且病程会相对延长并且会增加过早死亡的风险，同时长期的住院治疗使患者及其家庭需要承担不菲的费用。AN可归属于中医的“痞满”、“虚劳”、“纳呆”、“食少”等范畴。六君子汤源自明代虞抟编著的《医学正传》，该方由四君子汤加陈皮、半夏组成，具有健脾益气、燥湿化痰等功效，主治脾胃气虚兼痰湿症，食少便溏、胸脘痞闷、呕逆等。本文通过检索Pubmed、Web of science、知网等数据库，对六君子汤治疗AN国内外的临床报道进行了归纳总结。国医大师段富津运用六君子汤加味治疗因家长溺爱纵食所致厌食的3例患儿，取得显著疗效。周书兰等给120例AN患儿（平均年龄为6.7岁，男性患儿76例，女性患儿44例）以四逆散合六君子汤，并按照辨证施治的原则进行药物加减治疗，结果治愈62例，占51.7%；好转48例，占40.0%；无效10例，占8.3%，总有效率达91.7%。张娟娟等随机将AN患儿（平均年龄为3.42岁，男性患儿14例，女性患儿12例）分成对照组和治疗组，分别给予培菲康散（双歧杆菌三联活菌散）加硫酸锌口服液或六君子汤加减方进行治疗，发现培菲康散加六君子汤加减方组的总治疗效果更好，能够改善患者食欲，增加食量，并提高其血锌含量。金京丽以香砂六君子汤加白芍、枳实、山楂治疗1例15岁男性AN患者，其厌食症状得到有效缓解。日本学者报道，在对发病半年且未经治疗的5例AN患者（平均年龄21.2岁，均为女性）使用汉方六君子汤（Rikunshito）联合归脾汤进行治疗，全部病例症状均迅速改善，其中2例患者在短时间内明显好转。总之，六君子汤及其加减方在临床上治疗AN可取得满意疗效，可以有效地促进食欲、改善AN症状。但是，经检索文献发现，至今未有运用动物实验探究六君子汤治疗AN的作用机制的研究报道。鉴于六君子汤在临床上防治AN疗效确切，有必要对其作用机制进行深入研究。

关键词：六君子汤；神经性厌食

通讯作者：聂 克，E-mail：nicknk@hotmail.com

紫花杜鹃30-38馏分抗登革2型病毒的机制

晏丽君，刘俊珊，曹慧慧，区锦莹，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的体外实验评价紫花杜鹃30-38馏分（DJ 30-38）抗登革2型病毒（DENV-2）效果，并利用ICR乳小鼠模型进行抗病毒感染的验证。方法MTT法测药物对细胞毒性；CCK-8法检测药物对细胞的保护作用；结晶紫染色观察DJ 30-38不同浓度对DENV-2子代病毒释放的影响；实时荧光定量PCR检测不同浓度的DJ 30-38对DENV-2病毒复制中E和NS1蛋白基因表达的抑制率；Western印迹法检测不同浓度的DJ 30-38对DENV-2病毒中E和NS1蛋白表达的抑制。ICR乳小鼠分为空白、模型和DJ 30-38（5和10 mg·kg-1）4组，除空白组外的其余各组ICR乳小鼠颅内注射（400 PFU·mL-1）及ip给予（4×105PFU·mL-1）DENV-2病毒液20 μL，空白对照组注射等量RPMI-1640培养基，造模当天，DJ 30-38组乳小鼠提前2 h ig给药，以后每隔24 h给药1次，每天观察并记录各组乳小鼠的体重及发病情况，计算各组症状评分和生存率，取乳小鼠脑做病理切片的HE染色，用实时荧光定量PCR检测其他各脏器病毒E和NS1蛋白基因表达水平。结果DJ 30-38能显著改善DENV-2感染BHK-21细胞引起的细胞病变效应，抑制病毒子代的释放，抑制DENV-2感染后E和NS1基因表达水平，能显著降低病毒感染后E和NS1蛋白的表达。在DENV-2感染的ICR乳小鼠模型中，DJ 30-38改善了病毒感染乳小鼠的症状评分，延长了生存时间，给药各组乳小鼠脑、心、肝、脾、肺、肾等脏器的病毒拷贝数与模型组相比均降低，且呈剂量依赖性。结论DJ 30-38具有抗DENV-2的作用。

关键词：登革热；紫花杜鹃提取物；抗病毒；紫花杜鹃30-38馏分

通讯作者：余林中，E-mail：yulzh@smu.edu.cn

丹酚酸类药物SAA004和SAA008通过抑制NF- κB和STAT3信号通路发挥抗炎作用

杨 桦，邹丽芳，曹惠慧，余林中，卢子滨，区锦莹，刘俊珊

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的研究丹酚酸类化合物SAA004和SAA008的抗炎活性并探讨其潜在机制。方法采用卵黄囊显微注射内毒素（LPS）的炎症模型，通过生存分析观察SAA004和SAA008对注射LPS的斑马鱼的保护作用；通过H&E染色和荧光显微镜观察炎症细胞在卵黄囊的募集情况；通过q-PCR测量斑马鱼关键炎症细胞因子白细胞介素6（IL-6）和肿瘤坏死因子α（TNF-α）的转录水平；采用LPS诱导RAW264.7巨噬细胞建立炎症模型，通过ELISA检测RAW264.7巨噬细胞上清炎症因子IL-6和TNF-α的表达；通过Western印迹法检测RAW264.7巨噬细胞炎症相关信号通路（NF-κB和STAT3）中IκBα、STAT3的磷酸化及NF-κB-p65的表达；通过免疫荧光法检测RAW264.7巨噬细胞NF-κB-p65和p-STAT3的表达及分布。结果丹酚酸类药物SAA004和SAA008提高LPS刺激的斑马鱼生存率，减少LPS诱导的炎症细胞在卵黄囊部位的募集，降低炎症因子IL-6和TNF-α的转录水平；同时降低RAW264.7巨噬细胞中IL-6和TNF-α的表达水平，抑制IκBα和STAT3蛋白的磷酸化，抑制NF-κB-p65和p-STAT3蛋白的分布。结论丹酚酸类药物SAA004和SAA008通过抑制NF-κB和STAT3信号通路发挥抗炎作用。

关键词：SAA004；SAA00；斑马鱼；抗炎作用；NF-κB；STAT3

通讯作者：刘俊珊，E-mail：liujunshanty@163.com

痰热清注射液抗登革2型病毒的作用机制研究

杨家彬，刘俊珊，曹惠慧，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的探讨痰热清注射液抗登革2型病毒（DENV-2）的作用。方法不同稀释浓度的痰热清注射液作用感染DENV-2的huh-7，HepG2和MHCC97H肝癌细胞，实时荧光定量QPCR法检测huh-7细胞内病毒拷贝数；Western印迹实验检测huh-7细胞内病毒相关蛋白的表达量；病毒蚀斑实验检测子代病毒的释放量，RNA-seq比较宿主细胞内先天免疫通路相关基因的表达变化。结果不同稀释浓度的痰热清注射液均能抑制细胞内DENV-2的RNA和蛋白的复制，并减能少子代病毒的释放；RNA-seq结果显示细胞内多个先天免疫信号通路受DENV-2影响，且痰热清注射液能调控相关基因的表达。结论痰热清注射液具有抗DENV-2的作用。

关键词：痰热清注射液；登革病毒；RNA-seq

通讯作者：余林中，E-mail：yulzh@smu.edu.com

基于网络药理学探究丹皮酚治疗糖尿病心脏病作用

杨启帆1，邬东东1，徐 杰1，骆家引3，瞿皇皇4，赵 晶2

（陕西中医药大学1.公共卫生学院，2.国医大师研究院，3.第二临床医学院，4.护理学院，陕西 咸阳 712000）

摘要：目的探究丹皮酚（Pae）治疗糖尿病心脏病（DHD）的作用及机制。方法通过TCMSP数据库获得丹皮酚的ADME参数和理化参数；运用Pharm Mapper数据库预测丹皮酚的潜在靶点；采用DisGeNET数据库检索并构建DHD靶点数据库。筛选丹皮酚治疗DHD的潜在靶点，导入String数据库获得蛋白-蛋白直接相互作用网络，通过Cytoscape 3.8.2中Gene MANIA分析靶点共表达、物理相关性和遗传相关性等信息。利用DAVID数据库对其潜在靶点进行生物学过程、分子功能、细胞成分及信号通路分析，探讨丹皮酚治疗DHD的作用机制。采用软脂酸（PA）损伤心肌细胞构建DHD细胞模型，使用MTT细胞活力实验研究丹皮酚的保护作用，进一步运用JC-1实验观察丹皮酚对PA诱导的心肌细胞线粒体膜电位的影响，探究丹皮酚对DHD治疗作用及机制。结果通过TCMSP数据库分析得出丹皮酚口服利用度为28.79%，且理化参数符合里宾斯基药物筛选五规则。从PharmMapper数据库中得到丹皮酚潜在靶点54个，进一步分析获得丹皮酚治疗DHD潜在靶点14个。Gene MANIA数据库分析结果显示14个潜在靶点中共表达占48.87%、物理相关性占38.75%和遗传相关性占5.25%。对14个潜在靶点富集分析得出丹皮酚治疗DHD通过调控胰岛素抵抗、胰岛素信号通路和肿瘤坏死因子信号通路等信号通路，并参与蛋白水解作用、活性氧代谢过程和蛋白酪氨酸激酶活性的正调控和JAK-STAT级联调控等多种生物过程发挥对DHD的治疗作用。细胞活力实验结果显示与对照组相比，PA损伤组心肌细胞活力显著降低（P＜0.05）；与PA损伤组相比，丹皮酚干预组心肌细胞活力显著提高（P＜0.05），提示丹皮酚对PA损伤的心肌细胞具有保护作用。进一步研究发现，丹皮酚显著升高PA诱导的线粒体膜电位，表现在与PA损伤相比，丹皮酚干预后红色荧光强度显著增多，绿色荧光降低。结论丹皮酚对DHD有显著治疗作用，具有多靶点、多通路协同治疗作用，且其机制可能与影响线粒体膜代谢有关。

关键词：网络药理学；丹皮酚；糖尿病心脏病

基金项目：国家自然科学基金（82100488）；陕西省科技厅重点研发计划（2021SF-071）；国家级大学生创新创业训练计划（202110716027）

通讯作者：赵 晶，E-mail：zhaojing_1207@126.com

穿心莲内酯磺化物的抗炎活性评价

杨汤佳，刘俊珊，曹惠慧，区锦莹，余林中

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的利用斑马鱼炎症模型评价穿心莲内脂磺化物的抗炎活性。方法采用斑马鱼硫酸铜急性炎症模型，尾鳍横切局部炎症模型，LPS卵黄囊显微注射全身炎症模型评价穿心莲内酯磺化物的抗炎效果。结果FL-2，FL-3，FL-5，FL-10，FL-24和FL-33可显著抑制硫酸铜、尾鳍横切和内毒素诱导的斑马鱼中性粒细胞的迁移。结论FL-2，FL-3，FL-5，FL-10，FL-24和FL-33具有抗炎作用，提示其可以作为抗炎药物的候选药物。

关键词：炎症；斑马鱼模型；抗炎；穿心莲内脂磺化物

通讯作者：余林中，E-mail：yulzh@smu.edu.com

丹酚酸A对慢性脑缺血大鼠认知损伤及神经炎症的影响

杨玉娇1，2，宋俊科1，刘 珊1，3，张 森1，蒋 楠1，杨海光1，杜冠华1，2

（1.中国医学科学院北京协和医学院药物研究所，药物靶点研究与新药筛选北京市重点实验室，北京 100050；2.沈阳药科大学生命科学与生物制药学院，辽宁 沈阳 110017；3.广东药科大学中药学院，广东 广州 511436）

摘要：目的探讨丹酚酸A（SalA）对慢性脑缺血（CCI）大鼠认知损伤及神经炎症的影响。方法采用双侧颈总动脉结扎（2VO）法建立大鼠CCI模型，术后28 d采用Morris水迷宫（MWM）筛选模型，将造模成功大鼠随机分为2VO组，2VO+SalA（5，10和20 mg·kg-1）组，2VO+尼莫地平组，另设假手术组作为对照。通过MWM和旷场试验观察各组大鼠的认知功能，包括学习记忆能力和探索能力；采用HE染色、Nissl染色观察大鼠皮质和海马区的病理变化；通过酶联免疫吸附实验（ELISA）和免疫组化分析检测皮质和海马区炎症因子的表达和胶质细胞的活化。结果MWM测试显示，在空间探索实验中，SalA（5，10和20 mg·kg-1）明显缩短CCI大鼠潜伏期（P＜0.05），尼莫地平有缩短潜伏期的趋势但是效果不明显，SaLA 10和20 mg·kg-1组和尼莫地平组在60 s内穿越平台所在位置的次数明显增加（P＜0.05）。旷场测试显示，与模型组相比，2VO+NS组的运动总距离（P＜0.01）、站立次数（P＜0.01）和进入中心区域的频率（P＜0.05）显著降低。HE染色显示，正常神经元数目减少，可见神经细胞脱失，细胞形态不规则排列紊乱，出现神经细胞变性坏死，SalA可以改善这一现象，效果优于尼莫地平。Nissl染色显示，SalA 20 mg·kg-1和尼莫地平治疗可以显著减少CCI大鼠皮质神经元缺失（P＜0.05），并且SalA 10和20 mg·kg-1可以显著减少CCI大鼠海马神经元缺失（P＜0.05），而尼莫地平无显著作用。ELISA检测表明，SalA治疗明显降低了CCI大鼠皮质和海马区炎症因子TNF-α，IL-1β和IL-6的表达水平，而尼莫地平只对皮质区IL-6的表达具有明显降低的作用（P＜0.05）。免疫组化结果显示，经SalA治疗，CCI大鼠海马和皮质区GFAP、CD68的表达被明显降低（P＜0.05），而尼莫地平对2种蛋白的表达都没有影响。结论SalA可以通过降低炎症反应，减轻CCI引起的大鼠认知功能损伤。

关键词：慢性脑缺血；丹酚酸A；认知障碍；神经炎症

通讯作者：杜冠华，E-mail：dugh@imm.ac.cn

牛舌草总黄酮通过ROS/TXNIP/NLRP3抑制炎症小体活化抗心肌缺血再灌注损伤

阴苏月，姜 瑜，王丹姝，燕柳艳，王守宝，杜冠华

（中国医学科学院北京协和医学院药物研究所，药物靶点研究与新药筛选北京市重点实验室，北京 100050）

摘要：目的考察牛舌草总黄酮（TF）抗心肌缺血再灌注（I/R）损伤作用及其机制，探讨炎症小体活化和活性氧/硫氧还蛋白互作蛋白/NOD样受体蛋白3（ROS/TXNIP/NLRP3）通路是否参与其中。方法采用冠状动脉左前降支结扎制备小鼠I/R损伤模型。将模型制备成功的30只小鼠随机分为I/R模型组和I/R+TF（30和100 mg·kg-1）处理组，再灌同时给予生理盐水或TF。于再灌注24 h时，检测小鼠心功能，测定心肌梗死范围和血清心肌酶谱，心肌冰冻切片染色观察ROS水平，Western印迹法分析NLRP3炎症小体活化以及TXNIP与NLRP3相互作用的变化。结果TF 30和100 mg·kg-1均可明显改善心功能，减少心肌梗死范围，降低血清心肌酶水平。另外，TF处理减少心肌炎症因子IL-1β，IL-6和TNFα含量，降低心肌ROS水平。TF 30和100 mg·kg-1均显著降低TXNIP、NLRP3、活化的胱天蛋白酶1和活化的IL-1β，并抑制TXNIP与NLRP3相互作用。结论TF通过抑制ROS/TXNIP/NLRP3通路减轻NLRP3炎症小体活化，从而发挥抗心肌I/R损伤作用。

关键词：牛舌草总黄酮；心肌缺血再灌注损伤；NLRP3炎症小体；活性氧；TXNIP/NLRP3

基金项目：北京市自然科学基金（7192131）；国家自然科学基金（81673422）；国家科技重大专项（2018ZX09711001-010）

通讯作者：王守宝，E-mail：shoubaowang@imm.ac.cn；杜冠华，E-mail：dugh@imm.ac.cn

“中药化学生物学”驱动的天然活性分子抗脑缺血新靶点发现

曾克武

（北京大学药学院，天然药物及仿生药物国家重点实验室，北京 100191）

摘要：目的缺血性脑卒中是一类常见的脑血管病，具有高发病率、高死亡率的特点，目前尚缺少有效的药物治疗靶点。如何从中医药获得启发，利用其脑保护活性成分为化学工具探针，进而为缺血性脑中风新靶点的发现提供指导，这对于研发缺血性脑卒中的治疗药物具有重要的科学价值。方法基于“中药化学生物学”的指导思想，本研究选取了多个中药来源的抗脑缺血作用明确的天然活性分子作为化学探针，包括肉苁蓉的脑保护活性成分松果菊苷、苏木来源的原苏木素A、淫羊藿来源的淫羊藿苷等系列活性分子。通过靶点钩钓策略，从神经细胞中捕获其发挥脑保护作用的直接药理靶点，进而为缺血性脑卒中的临床治疗及创新药物研发提供源头靶点信息。结果发现松果菊苷的直接神经保护靶点为酪氨酸激酶的a′催化亚基（CK2α′），即松果菊苷靶向结合CK2α′，并通过非经典激酶方式活化Wnt/β-catenin信号通路，诱导线粒体融合并发挥抗脑缺血作用。同时还发现，原苏木素A通过作用于14-3-3ζ蛋白发挥抗脑缺血作用，淫羊藿苷通过作用于Bax蛋白发挥脑保护作用。结论这些工作阐明了松果菊苷等系列中药活性成分的直接脑保护靶点，为今后治疗缺血性脑卒中的创新药物设计与研发提供了关键的靶点生物学信息。

关键词：缺血性脑卒中；中药化学生物学；中药活性分子探针；靶点钩钓；肉苁蓉

通讯作者：曾克武，E-mail：ZKW@bjmu.edu.cn

作用于细菌群体感应系统的抗菌中药单体

曾 利1，凌保东1，2

（成都医学院1.药学院，2.结构特异性小分子药物研究四川省高校重点实验室，四川 成都 610500）

摘要：近年来，很多天然药物已经分离出了多种抗菌单体化合物可以影响细菌群体感应系统，调节相关毒力因子的表达，或通过与抗菌药物联合使用，增强抗菌药物的抗菌作用效果。本文主要对作用于细菌群体感应系统的中药单体抗感染作用研究进展进行综述。1.群体感应是一种细菌间通信交流机制，细菌通过自身分泌的信号分子感知自身周围群体密度，其信号分子随着群体密度的增加而增加，当达到一定的阈值时，启动菌体中特定基因的表达，从而导致多种细菌行为。细菌群体感应系统主要分为3类：①在革兰阴性菌中由酰基高丝氨酸内酯（AI-1）介导；②在革兰阳性菌中的由自身诱导肽（AIP）介导；③在革兰阴性菌和革兰氏阳性菌中均能由硼酸二酯（AI-2）介导。干扰群体感应系统可影响细菌生物被膜的形成和毒力因子的分泌以及外排泵的表达等，这为临床耐药菌的治疗和生物被膜菌感染的防控提供了新的思路。2.具有群体感应抑制作用的中药单体有黄酮类、苯甲酸衍生物、萜类化合物、香豆素类、生物碱类等，都具有抗群体感应的特性，并且主要通过抑制信号分子的合成、促进信号分子的降解和与信号分子竞争结合受体蛋白3种机制发挥作用。3.将中药单体作为群体感应抑制剂与传统抗菌药物结合是提高抗菌药物的抗菌效果以及恢复抗菌药物对细菌敏感性的一种新的策略。越来越多的抗菌药物包括头孢吡肟、头孢他啶和亚胺培南等发现具有群体感应抑制活性，黄芩苷联合替加环素对泛耐药鲍曼不动杆菌能够产生协同抗菌作用，姜黄素和头孢他啶联合使用能够恢复抗菌药物对细菌的敏感性。这些中药单体作为群体感应抑制剂与其他群体感应抑制剂或抗菌药物联合使用可以出现协同或相加的抗菌效果。中药单体作为群体感应抑制剂作与抗菌剂联合应用于耐药细菌，通过影响细菌的群体感应系统，不仅可产生协同抗菌和协同抗生物被膜作用，还能有效降低细菌的耐药性形成，为临床多重耐药菌以及生物被膜菌引起的感染提供了新的治疗方案。

关键词：细菌群体感应系统；中药单体；抗菌

基金项目：国家自然科学基金（81373454）

通讯作者：凌保东，E-mail：lingbaodong@cmc.edu.cn

六味地黄丸对顺铂所致遗传及生殖毒性的改善作用及机制

张 阔，杨静玉，吴春福

（沈阳药科大学药理教研室，辽宁 沈阳 110016）

摘要：目的顺铂作为临床经典化疗药物，对多种肿瘤均有较好的治疗效果，但其严重的消化系统、造血系统、肾、遗传及生殖毒性成为其临床应用受限的主要因素。近年来，化疗引起的生殖系统损伤越来越受到关注，但临床治疗药物匮乏。有研究表明,六味地黄方剂可改善顺铂所致的肾毒性，且六味地黄方剂与顺铂联用可增强顺铂的抗肿瘤效果，但六味地黄方剂对顺铂所致遗传、生殖毒性是否具有保护作用，尚未见相关报道。本文研究六味地黄丸对顺铂所致小鼠遗传及生殖毒性的改善作用及其氧化应激相关机制。方法采用单细胞凝胶电泳技术、微核试验、吖啶橙染色、伊红染色、精子计数法、精子畸形实验和睾丸HE染色考察了六味地黄丸对顺铂诱导的小鼠外周血细胞DNA损伤、微核生成、精子DNA完整率下降、精子存活率降低、精子数量下降、精子畸形率升高和睾丸细胞病变的保护作用。结果六味地黄丸改善顺铂对小鼠外周血细胞DNA的损伤，改善顺铂所致小鼠骨髓细胞微核生成的抑制，但未能改善由顺铂导致的小鼠体重及睾丸指数的变化；六味地黄丸增加顺铂诱导小鼠的精子DNA完整率、精子存活率及精子数量，降低顺铂诱导小鼠的精子畸形率，但不影响其畸形类型的构成；六味地黄丸改善顺铂诱导小鼠的睾丸组织病变；六味地黄丸降低顺铂诱导小鼠睾丸组织的MDA水平，提高顺铂诱导小鼠睾丸组织的T-SOD、CAT、GSH-Px活性。结论六味地黄丸连续ig给药14 d对顺铂所致遗传及生殖毒性有改善作用，机制可能与改善顺铂所致氧化应激有关。

关键词：顺铂；六味地黄丸；遗传毒性；生殖毒性

通讯作者：吴春福，E-mail：wucf@syphu.edu.cn

帕金森病治疗药物研究进展

张程璐，陈乃宏，苑玉和

（天然药物活性物质与功能国家重点实验室，中国医学科学院药物研究所，北京 100050）

摘要：帕金森病（PD）是一种神经退行性疾病，其致病机制尚未完全阐明。目前采用的PD治疗药物主要为多巴胺能药物，如美多芭等，其通过补充脑内多巴胺的含量来缓解静止性震颤、肌强直等症状。但多巴胺能药物仅能做到对症治疗，并不能延缓PD的病程发展，同时其使用也受到一些不良反应的限制。目前研发PD治疗药物的研发策略主要是改进已有的多巴胺能药物、针对最新研究发现的新的分子靶标开发新药和通过“老药新用”发掘潜在的治疗药物。其中，大部分的研究关注于新的分子靶标，包括烟酰胺腺嘌呤二核苷酸磷酸酯（NADPH）氧化酶、核受体相关1（Nurr1）、腺苷受体A2、尼古丁受体、代谢性谷氨酸受体（mGluRs）和葡萄糖脑苷酶（GCase）等，并取得了较好的研究进展，一些新型化合物已在动物实验及临床水平上展现出了良好的治疗效果。另外，最新研究表明PD与阿尔茨海默病和糖尿病具有部分相同的病理改变，因此研究也关注治疗阿尔茨海默病和糖尿病的药物能否用于PD的治疗。同时，针对α-突触核蛋白的抗体在动物实验中几乎能够完全清除错误的α-突触核蛋白，提示免疫疗法可能成为治疗PD的有效手段。综上所述，研究通过改进多巴胺能药物、开发新型化合物、以及“老药新用”等策略，PD的治疗药物取得了良好的研究进展，并为未来的研究提示了发展方向。

关键词：帕金森病；多巴胺能药物；分子靶标；α-突触核蛋白

通讯作者：苑玉和，E-mail：yuanyuhe@imm.ac.cn

西藏凹乳芹化学成分及药理作用研究进展

张梦如，赵健黎，孙芳云

（西藏民族大学医学部，西藏民族大学藏药检测技术教育部工程研究中心，陕西 咸阳 712082）

摘要：西藏凹乳芹（Vicatia thibeticade Boiss）是藏药“加哇”的汉译名，为伞形科凹乳芹属植物凹乳芹的根，在云南称其为“西归”，是常用的药食两用藏药，为藏药五大根药之一。其性温、味辛、甘、微苦，具有祛风除湿、散寒止痛的作用。西藏凹乳芹在全世界大约有4种，产于中国、印度、巴基斯坦和尼泊尔，在我国主要分布于西藏、四川、云南等地。主要化学成分为伞形花内酯、佛手苷内酯、β-谷甾醇、胡萝卜甙、氨基酸、黄酮、多糖、芹菜素和阿魏酸等。毒理研究表明，西藏凹乳芹根对血液生化指标、脏器指数、体重及摄食量无影响，长期服用未发现毒性反应，属于无毒级，有较高的食用安全性。西藏凹乳芹药理作用研究多集中在抗疲劳、抗氧化和抗衰老等方面。其药理机制主要有①抗疲劳：西藏凹乳芹根可明显增加体内红细胞数和血红蛋白含量，增加肌肉中肌糖原含量，减少乳酸和丙二醛的含量，降低乳酸脱氢酶活性，提高超氧化物歧化酶活性，进而提高机体抗疲劳作用。②抗氧化：西藏凹乳芹根中的黄酮能较好地清除羟自由基、超氧阴离子自由基和DPPH自由基以提高抗氧化能力。③抗衰老：西藏凹乳芹根可以通过激活DNA损伤修复途径修复DNA的损伤，从而减轻衰老细胞中的周期停滞和肝损伤。但西藏凹乳芹在抗疲劳、抗氧化、抗衰老等方面的具体化学成分尚不清楚，其它现代药理学研究报道很少，开发应用不够，有待于进一步研究。

关键词：西藏凹乳芹；药理；毒理；化学成分

基金项目：2020年改革发展专项——藏药检测技术教育部工程研究中心（324042000101）

通讯作者：孙芳云，E-mail：xzmysfy@163.com

TLR4-IN-C34通过抑制TLR4/MyD88/NF- κB/NLRP3信号通路减轻脂多糖诱导的BV2小胶质细胞炎症反应

张姗姗，刘 漫，刘冬妮，杨滢霖，王月华，杜冠华

（药物靶点研究与新药筛选北京市重点实验室，中国医学科学院北京协和医学院药物研究所，北京 100050）

摘要：目的研究TLR4-IN-C34在LPS刺激的BV2小胶质细胞中的抗炎作用及其可能机制。方法LPS刺激BV2细胞建立细胞水平神经炎症模型；Griess反应测NO水平；ELISA检测促炎细胞因子水平；Western印迹法和RT-PCR检测了iNOS和COX-2蛋白以及与TLR4/MyD88/NF-κB/NLRP3信号通路相关的几种蛋白的转录和表达水平的影响；流式细胞术检测ROS水平。结果在LPS诱导BV2细胞模型上，TLR4-IN-C34抑制了LPS刺激的BV2细胞中NO的产生、促炎因子包括TNF-α，IL-1β，IL-6和MCP-1的水平以及iNOS和COX-2的转录和表达水平。TLR4-IN-C34显著降低了TLR4、MyD88、NLRP3、活化胱天蛋白酶1、IL-18和IL-1β的表达水平以及NF-κB和IκBα蛋白的磷酸化水平。此外，TLR4-IN-C34在LPS处理后显著降低了BV2细胞的ROS水平，表现出强烈的抗氧化作用。结论在LPS刺激的BV2细胞中，TLR4-IN-C34减少促炎因子水平，表现出抗炎活性，其机制可能与抑制TLR4/MyD88/NF-κB/NLRP3信号通路的激活有关。

关键词：TLR4-IN-C34；神经炎症；Toll受体4；BV2小胶质细胞

基金项目：国家重点研发计划（2018YFC0311005）；国家科技“十三五”重大专项（2018ZX09711001-003-019）；中国医学科学院医学与健康科技创新工程（2016-I2M-3-007）

通讯作者：王月华，E-mail：walling@imm.ac.cn；杜冠华，E-mail：dugh@imm.ac.cn

天麻素对小鼠酒精性肝损伤的保护作用

张逍遥1，李 奔1，李鑫鑫1，陈 琛1，张小莺1，2

〔1.陕西理工大学生物科学与工程学院中德天然产物研究联合实验室，陕西汉中 723000；2.Centre of Molecular and Environmental Biology（CBMA），Department of Biology，University of Minho，Braga 4710-057，Portugal〕

摘要：目的探讨天麻素对酒精性肝损伤的保护作用。方法对小鼠进行酒精肝毒性实验，并用天麻素（50，80和100 mg·kg-1，ig）治疗。血清谷丙转氨酶（GPT）、谷草转氨酶（GOT）、超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GPx）和过氧化氢酶（CAT）的活性以及丙二醛（MDA）水平用商业诊断试剂盒进行检测。对肝组织进行切片分析。利用Western印迹法分析微粒体组分中细胞色素P450 2E1酶（CYP2E1）的表达。利用实时荧光定量PCR系统检测该蛋白的mRNA的表达情况。结果天麻素50，80和100 mg·kg-1组小鼠的GOT和GPT酶的活性和肝中MDA的含量低于模型组小鼠，而SOD，GPx和CAT的活性高于模型组小鼠，且呈剂量依赖性；肝组织切片病变情况优于模型组；CYP2E1的表达水平也低于模型组小鼠。结论天麻素能够有效降低酒精引起的GPT、GOT和CYP2E1酶活性的增强以及肝组织MDA含量的升高，增强抗氧化酶SOD，GPx和CAT的活性，缓解酒精引起的肝组织病变，从而改善小鼠酒精性肝损伤。

关键词：天麻；酒精；肝损伤；抗氧化剂；炎症

通讯作者：张小莺，E-mail：zhang@bio.uminho.pt

富含α-亚麻酸的亚麻籽油通过调节肠道菌群和抑制炎症改善大鼠2型糖尿病

张晓霞，穆 静，杨正飞

（宁夏医科大学中医学院，宁夏 银川 750004）

摘要：目的评价富含α-亚麻酸（ALA）的膳食亚麻籽油（FO）对2型糖尿病（T2DM）大鼠的干预效果与机制。方法将SD大鼠随机分为4组（每组10只）：正常对照组（PF/CO）、T2DM模型组（DM/CO）、FO对照组（PF/FO）和FO干预组（DM/FO）SD大鼠禁食过夜，依次ip给予链脲佐菌素（STZ）和烟酰胺（NA）溶液，成功诱导T2DM模型。膳食FO干预5周，每周定期监测大鼠体重（BW）和空腹血糖（FBG）。干预结束后，收集大鼠粪便，通过16S rRNA高通量测序，分析不同组大鼠肠道菌群多样性及丰度的变化；利用气相色谱-质谱（GC-MS）检测每组大鼠粪便中短链脂肪酸（SCFA）含量的变化。在FO干预5周后麻醉处死大鼠，收集血浆（清）及组织样本，采用ELISA试剂盒检测大鼠血浆糖化血红蛋白（GHb）、胰岛素（INS）、炎症因子白细胞介素（IL）-6、IL-17A、IL-1β、肿瘤坏死因子（TNF）-α和IL-10的浓度；显色基质鲎试剂盒检测SD大鼠血浆中脂多糖（LPS）的水平；Spearman相关性分析血浆炎症因子与肠道菌群相关性，Pearson相关性分析炎症因子与SCFAs相关性。结果DM/CO组大鼠BW较PF/CO组明显降低，而FBG、GHb及血脂水平显著升高，FO干预显著改善T2DM大鼠BW，FBG和GHb水平。DM/CO组血浆炎症指标（IL-1β，IL-6，IL-17A，TNF-α和LPS）较PF/CO组明显增高，FO干预后T2DM大鼠炎症指标较DM/CO组显著降低，但抗炎因子IL-10水平在各组间无统计学差异。肠菌测序显示，DM/CO组与PF/CO组相比，在门水平上，Firmicutes（厚壁菌门）相对丰度明显升高，Bacteroidetes（拟杆菌门）显著降低，在属水平上，Blautia（布劳特氏菌）显著升高，而且Alistipes（另枝菌）含量明显降低。然而，与DM/CO组相比，DM/FO组Firmicutes和Blautia相对丰度显著降低，Tyzzerella也降低，但无统计学意义，而Bacteroidetes和Alistipes含量明显升高。Spearman相关分析显示：Firmicutes和Blautia与血浆炎症因子（IL-1β，TNF-α，IL-6和LPS）成正相关，Bacteroidetes和Alistipes与炎症因子（TNF-α和LPS）成负相关。GS-MS结果显示，DM/CO组与PF/CO组相比，SCFA（乙酸，丙酸和丁酸）含量降低，但丙酸和丁酸在2组间无显著差异；而FO干预后，DM/FO组乙酸、丙酸和丁酸含量显著升高。Pearson相关性分析发现，乙酸、丙酸、丁酸分别与炎症因子（IL-1β，IL-6和IL-17A）成负相关。结论富含ALA的膳食FO可通过抑制炎症反应、调节肠道菌群及代谢产物SCFA缓解大鼠T2DM。

关键词：α-亚麻酸；糖尿病；炎症；肠道菌群；短链脂肪酸

通讯作者：张晓霞，E-mail：zxx1216@163.com

女贞苷抑制RIPK1/RIPK3/MLKL通路减轻脑卒中大鼠神经元坏死样凋亡

张议月1，刘 斌2，罗秀菊3，彭 军1

（1.中南大学湘雅药学院药理学系，心血管研究湖南省重点实验室，湖南 长沙 410078；2.中南大学湘雅医院药学部，湖南 长沙 410008；3.中南大学湘雅三医院检验科，湖南 长沙 410013）

摘要：目的探究女贞苷对脑卒中大鼠脑损伤的影响及其作用机制。方法采用大脑中动脉闭塞（MCAO）法建立缺血性脑中风大鼠模型，Longa评分法评估大鼠神经功能，TTC染色检验大鼠脑梗死体积，TUNEL染色检测大鼠脑组织细胞凋亡，MTS法检测PC12细胞活力，流式细胞术检测PC12细胞坏死样凋亡，Western印迹法检测大鼠脑组织和PC12细胞RIPK1，RIPK3，MLKL和p-MLKL蛋白表达，免疫共沉淀检测PC12细胞中RIPK3与RIPK1和MLKL之间的相互作用，Molecular Operating Environment（MOE）软件分析女贞苷与RIPK1，RIPK3和MLKL之间的相互作用。结果大鼠脑缺血2 h再灌注24 h后，与假手术组相比，缺血/再灌组神经功能学评分、脑梗死体积增加，RIPK1，RIPK3，MLKL和p-MLKL蛋白表达上调，胱天蛋白酶3活性增强；女贞苷30 mg·kg-1可降低缺血大鼠神经功能学评分和脑梗死体积，下调RIPK3，MLK和p-MLK蛋白表达，减少脑细胞凋亡和胱天蛋白酶3活性。PC12细胞低氧8 h复氧24 h后，与常氧组相比，低氧组细胞活力降低，LDH释放（～25%）、细胞坏死样凋亡（～17%）增多，RIPK1，RIPK3，MLKL和p-MLKL蛋白表达上调，RIPK3与RIPK1和MLKL相互作用增强；女贞苷可增加低氧细胞活力，降低LDH释放（～13%）、细胞坏死样凋亡（～7%），下调RIPK3，MLKL和p-MLKL蛋白表达，抑制RIPK3与RIPK1和MLKL相互作用；MOE软件分析结果显示，女贞苷可能与RIPK1 24位Asp、142位Glu，RIPK3 32位Gly、33位Phe、142位Arg、193位Leu和MLKL 21位Met、25位Lys、154位Asp存在相互作用。结论女贞苷通过靶向抑制RIPK1/RIPK3/MLKL通路减轻脑卒中大鼠神经元坏死样凋亡。

关键词：女贞苷；坏死样凋亡；脑卒中；RIPK1；MLKL

通讯作者：彭 军，E-mail：jungpeng@csu.edu.cn；罗秀菊，E-mail：xjluo22@csu.edu.cn

丹参素对血管内皮细胞氧化损伤保护作用机制研究进展

张钰炜，张丹参，景永帅

（河北科技大学化学与制药工程学院，河北 石家庄 050018）

摘要：随着人们生活水平的不断提高，心血管系统疾病发病率逐年升高。研究发现，动脉粥样硬化是常见的心血管系统发病原因，而肥胖、老龄化、紧张状态等都会引起动脉粥样硬化。血管内皮细胞氧化损伤是动脉粥样硬化的始动因素，并伴随着血栓、冠心病等疾病的发生。因此，保护氧化损伤后的血管内皮细胞是预防和治疗心血管疾病的重要方法，利用天然化学成分保护受损血管内皮细胞是近年来的研究热点。丹参是我国传统的药食同源药材，具活血祛瘀、通经止痛功效，包含丹参素、丹参酮和丹酚酸等多种有效成分。其中丹参素具有抗氧化应激、扩张冠脉血管、抑制血小板聚集等药理活性，是保护血管内皮细胞氧化损伤的一种有效的天然药物。丹参素对血管内皮细胞氧化损伤保护的作用机制主要包括：①抑制细胞凋亡：丹参素可以抑制线粒体膜电位的下降，提高抗凋亡蛋白Bcl-2的表达，降低促凋亡蛋白Bax和肿瘤抑制蛋白P53的表达，抑制促凋亡蛋白胱天蛋白酶9和胱天蛋白酶3的活化和DNA修复酶PARP的断裂，从而抑制细胞凋亡。此外，丹参素可以通过抑制内皮细胞内p38以及JNK信号通路抑制内皮细胞的凋亡。②抑制活性（ROS）氧的产生：丹参素可剂量依赖性地抑制活性氧所致细胞活力降低，通过改变血管内皮细胞的线粒体膜电位，显著减少丙二醛（MDA）生成，提高超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）活力，一氧化氮含量（NO）和内皮型一氧化氮合成酶（eNOS）的表达，起到对ROS损伤内皮细胞的保护作用。③抗炎作用：丹参素可减少炎症因子白细胞介素6（IL-6）和肿瘤坏死因子α（TNF-α）的分泌，同时抑制内皮细胞内皮素1（ET-1）mRNA的表达，升高过氧化物酶体增生物激活受体γ（PPAR-γ）mRNA，血栓调节蛋白（TM）mRNA的表达，从而减轻炎症反应，减少氧化损伤。④抗凝血作用：丹参素可提高红细胞的负电荷，减少红细胞团聚，降低血液粘度，从而抑制血小板聚集。另外，丹参素可激活纤溶酶系统，增加组织纤维蛋白溶酶原激活剂（TPA）蛋白表达，减少纤溶酶原激活物抑制剂1（PAI-1）蛋白表达，从而发挥抗凝作用。综上，丹参素能够有效治疗和保护血管内皮细胞氧化损伤，在医药和保健品领域存在着巨大的开发潜力。

关键词：丹参素；血管内皮细胞；氧化损伤

基金项目：河北省省级科技计划（H2021208007）

通讯作者：景永帅，E-mail：cjys1985@126.com

姜黄素抑制纤维蛋白肽诱导血管内皮细胞炎症反应的作用机制

赵 晶1，2，徐守竹3，刘香柠1，马萌萌1，屈 伸1，乔海法1，2

（陕西中医药大学1.针灸推拿学院，2.国医大师研究院，3.公共卫生学院，陕西 咸阳 712000）

摘要：目的探讨姜黄素（CUR）对纤维蛋白肽A（FPA）诱导血管内皮细胞炎症反应的作用机制。方法①体外实验：培养人脐静脉血管内皮细胞（HUVEC），筛选CUR对FPA诱导HUVEC功能障碍的最佳药物浓度，采用CCK-8检测细胞活力，透射电镜观察细胞形态，激光共聚焦显微镜观察活性氧（ROS）水平，ELISA法检测上清中IL-6，IL-1β和CRP炎症因子的含量，免疫荧光检测核因子κB（NF-κB）的活化，Western印迹法检测MAPK信号通路蛋白分子的表达。②体内实验：大鼠体内微渗透泵复制高FPA血症大鼠模型，采用ELISA试剂盒检测血浆中FPA，IL-6，IL-1β和CRP的含量，免疫荧光检测血管内皮脱落情况以及ROS的表达情况，血管肌动扫描实验检测血管内皮功能，Western印迹法检测MAPK，NF-κB信号通路蛋白分子的表达。结果①体外实验：姜黄素预处理HUVEC 1 h后，CUR 5，25和50 μmol·L-1组细胞活力无影响；与FPA组比较，CUR组细胞较完整，空泡较少，线粒体形态较好，线粒体膜电位有明显升高作用，ROS表达水平显著降低（P＜0.05），p-ERK1/2和p-P38蛋白表达量显著下调（P＜0.05），NF-κB活化水平明显降低。②体内实验：CUR组干预后能够改善血管内皮功能障碍及内皮脱落情况，血浆中FPA，IL-6，IL-1β和CRP的含量显著降低（P＜0.05），血管ROS的表达降低，p-ERK1/2、p-P38和NF-κB蛋白表达量显著下调（P＜0.05）。结论姜黄素具有抑制FPA诱导血管内皮细胞炎症反应的作用，其机制与调节ROS-MAPK-NF-κB通路有关。

关键词：姜黄素；纤维蛋白肽A；血管内皮细胞；炎症反应

基金项目：国家自然科学基金（821005016）；陕西省科技厅重点研发计划（2021SF-071）；国家级大学生创新创业训练计划（202110716027）

通讯作者：乔海法，E-mail：1805355963@qq.com

基于网络药理学方法探讨连翘防治化疗性恶心呕吐的作用机制

赵耀中，聂 克

（广东药科大学中药学院，广东 广州 510006）

摘要：目的采用网络药理学方法预测分析连翘防治化疗性恶心呕吐（CINV）的潜在靶点，以进一步探讨连翘的止呕作用机制。方法利用TCMSP数据库按口服生物利用度（OB）≥30%和类药性（DL）≥0.18、BANTMAN-TCM数据库按评分≥20为筛选条件、结合文献筛选连翘的有效成分；利用PubChem数据库、ALOGPS 2.1在线软件和Swiss Target Prediction数据库收集连翘有效成分的治疗靶点；利用TTD数据库、GeneCards数据库和DrugBank数据库收集CINV的疾病靶点；利用STRING数据库构建蛋白-蛋白相互作用（PPI）网络，分析连翘有效成分防治CINV靶点之间的关系；利用DAVID6.8数据库进行GO分析和KEGG分析；利用cytoscape3.8.2软件绘制“连翘有效成分-CINV靶点-信号通路网络”并进行网络拓扑参数分析。结果获得汉黄芩素（wogonin）、牛蒡苷（arctiin）、山奈酚（kaempferol）、连翘苷（phillyrin）和连翘酯苷A（forsythoside A）等31个连翘有效成分；筛选得到丝氨酸/苏氨酸蛋白激酶1（Akt1）、天冬氨酸特异性的半胱氨酸蛋白酶3（CASP3）、表皮生长因子受体（EGFR）、5-羟色胺3A受体（5-HTR3A）、酪氨酸蛋白激酶2（JAK2）、单胺氧化酶A（MAO-A）等87个连翘防治CINV的潜在作用靶点；在GO分析中，连翘防治CINV共涉及344个生物学过程、56个细胞组分和83个分子功能，生物学过程主要涉及肽基酪氨酸磷酸化、蛋白质磷酸化、对药物的反应等，细胞组分主要涉及质膜、受体复合物、膜筏、蛋白质复合物等，分子功能主要涉及跨膜受体蛋白酪氨酸激酶活性、ATP结合、蛋白酪氨酸激酶活性等；KEGG分析筛选得到113条连翘防治CINV的信号通路，主要涉及PI3K-Akt信号通路、Rap1信号通路、Ras信号通路等。结论连翘防治CINV具有多成分、多靶点、多环节综合作用的特点，干预PI3K-Akt信号通路、Rap1信号通路及Ras信号通路可能是连翘防治CINV的潜在作用靶点。

关键词：连翘；化疗性恶心呕吐；网络药理学

基金项目：广东省重点学科科研项目（2019-GDXK-0011）

通讯作者：聂 克，E-mail：nicknk@hotmail.com

定痫丸中抗癫痫药对的再开发研究

赵泽丰

（陕西中医药大学陕西省针药结合重点实验室，陕西 咸阳 712046）

摘要：目的为从中药经方中发现抗癫痫临床候选药物提供理论依据。方法根据中医“君臣佐使，七情和合”的配伍理论，课题组前期对定痫丸进行了拆方研究，定痫丸为中医治疗痫证的重要方剂，载于清代《医学心悟》，用于治疗癫痫的突然发作，拆方后得到了对癫痫具有治疗作用的“远志-天麻”，“远志-石菖蒲”等药对。方中远志养心安神，天麻熄风止痉，石菖蒲开窍化痰。课题组结合网络药理学、虚拟筛选、亲和超滤、结构修饰、整体动物（MES，PTZ，3-MP）、分子病理（WB，HE，IP）及电生理等手段对上述药对治疗癫痫的机理进行了探讨。结果①物质基础：采用液质联用手段对“远志-天麻”提取物进行成分鉴定，结合文献报道建立“远志-天麻”成分化合物库。在此基础上构建“成分-靶点-通路”网络模型，预测远志与天麻治疗癫痫的有效成分及潜在作用机制。②虚拟筛选：通过文献报道的GABA-AT抑制剂为训练集提取其化学特征信息构建了基于配体的HipHop药效团，筛选“远志-天麻”中与药效团匹配的成分，采用分子对接及分子动力学模拟等手段对满足GABA-AT药效团要求的成分与靶标蛋白作用进行了评估，结果表明远志中的细叶远志皂苷A及西伯利亚远志糖A5与天麻中的天麻素等成分可能具有较好的GABA-AT抑制作用。③结构修饰：我们先后对远志活性成分TMCA衍生物，石菖蒲活性成分细辛醚衍生物及远志不同部位提取物抗癫痫活性进行系统评价。以对药“远志-天麻”的代谢效应成分TMCA及香兰醇为先导物进行结构修饰，合成TMCA-天麻苷元酯、TMCA-香兰酯、TMCA-乙酰天麻素酯等衍生物衍生物并进行了抗癫痫活性测定，结果表明多种衍生物的保护指数远高于阳性药物卡马西平，其中代表性化合物TMCA-乙酰天麻素酯（ED50：19.5 mg·kg-1）具有进一步开发的潜在价值。④分子病理及电生理：初步研究结果表明“远志-天麻”提取物能够对癫痫状态下的焦虑抑郁样行为起到一定治疗作用，调控脑内海马区GABA-AT/GAT-1/GAD 65/GABRA1轴蛋白的表达水平，减少神经元脱失，降低CA1区及CA3区GAT-1，GABA-AT及GABRA1免疫阳性细胞数，而“远志-石菖蒲”活性成分则能够作为乳酸脱氢酶/GABAAR双靶标活性化合物，显著促进氯离子内流，形成神经细胞膜超极化状态。结论基于中药经方定痫丸的药对的再开发研究能够为中药新要的发现及癫痫的中西医结合治疗提供科学依据。

关键词：定痫丸；癫痫；网络药理学；虚拟筛选；结构修饰

基金项目：陕西中医药大学经脉-脏腑相关研究创新团队（YL-09）；国家自然科学基金（8210142829）

作者简介：赵泽丰，E-mail：zzf598155752@sina.com

联苄基化合物20C通过调节α-突触核蛋白相关的炎症反应保护MPTP/P致小鼠毒性

周诗绮，张秋双，苑玉和，陈乃宏

（天然药物活性物质与功能国家重点实验室，中国医学科学院药物研究所&神经科学中心，北京 100050）

摘要：目的探究联苄基化合物20C在帕金森病（PD）小鼠体内的保护作用。方法通过1-甲基-4-苯基-1，2，3，6-四氢吡啶/丙磺舒（MPTP/p）小鼠模型，应用爬杆测试评价小鼠的运动能力；通过高效液相色谱（HPLC）检测20C对小鼠脑内多巴胺（DA）含量和代谢的影响；应用免疫组织化学和Western印迹法评价20C对小鼠黑质和纹状体神经元数量和活性的影响以及对小鼠脑内α-突触核蛋白（α-syn）及其聚集态、硝化和磷酸化形式含量变化的影响；通过透射电镜观察该化合物对小鼠黑质超微结构的影响；应用免疫荧光化学检测20C对黑质小胶质细胞和星形胶质细胞的影响。结果20C改善行为缺陷，减少DA消耗，减少多巴胺能神经元丢失，保护血脑屏障结构，改善突触核蛋白功能障碍，抑制胶质细胞活化，调节NF-κB信号和NOD样受体蛋白（NLRP）3炎症小体途径。结论20C通过减少突触核蛋白功能障碍和调节突触核蛋白相关的炎症反应对MPTP/p模型小鼠发挥保护作用。

关键词：帕金森病；α-突触核蛋白；黑质；多巴胺神经元

通讯作者：苑玉和，E-mail：yuanyuhe@imm.ac.cn；陈乃宏，E-mail chennh@imm.ac.cn

基于斑马鱼模型的康复新液药效学评价

邹丽芳，曹惠慧，余林中，区锦莹，刘俊珊

（南方医科大学中医药学院中药药理教研室，广东 广州 510515）

摘要：目的利用斑马鱼人类疾病模型评价康复新液的促进血管新生、修复组织及抗炎作用。方法采用索拉菲尼诱导的节间血管缺陷模型评价康复新液促进斑马鱼血管生成的作用；通过斑马鱼的尾鳍横切模型评价康复新液促进尾鳍再生的作用；通过斑马鱼硫酸铜、尾鳍横切及卵黄囊显微注射内毒素的炎症模型评价康复新液的抗炎作用，并采用实时荧光定量PCR法检测康复新液对炎症因子白细胞介素6（IL-6）和肿瘤坏死因子-α（TNF-α）mRNA表达水平的影响。结果康复新液能提高斑马鱼的节间血管指数；增加斑马鱼横切尾鳍的再生长度；抑制硫酸铜、鱼尾鳍横切和内毒素诱导的斑马鱼中性粒细胞的迁移，降低内毒素刺激的斑马鱼IL-6和TNF-α mRNA的表达水平。结论康复新液具有促进斑马鱼血管新生、促进组织修复和抗炎的作用。

关键词：康复新液；斑马鱼；炎症；血管新生；组织修复

通讯作者：刘俊珊，E-mail：liujunshanty@163.com