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Article(id=1322923593239232874, tenantId=1146029695717560320, journalId=1255847919539208197, issueId=1322923268038078806, articleNumber=null, orderNo=null, doi=10.13193/j.issn.1673-7717.2025.04.001, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=null, receivedDateStr=null, revisedDate=null, revisedDateStr=null, acceptedDate=null, acceptedDateStr=null, onlineDate=1793339163359, onlineDateStr=2026-10-30, pubDate=1744214400000, pubDateStr=2025-04-10, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1793339163359, onlineIssueDateStr=2026-10-30, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1793339163359, creator=13701087609, updateTime=1793339163359, updator=13701087609, issue=Issue{id=1322923268038078806, tenantId=1146029695717560320, journalId=1255847919539208197, year='2025', volume='43', issue='4', pageStart='1', pageEnd='258', issueExtLink='null', onlineDate='null', pubDate='1744214400000', pubDateStr='2025-04-10', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=0, createTime=1793339085826, creator='13701087609', updateTime=1793340586453, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1322929562572686061, tenantId=1146029695717560320, journalId=1255847919539208197, issueId=1322923268038078806, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1322929562572686062, tenantId=1146029695717560320, journalId=1255847919539208197, issueId=1322923268038078806, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=1, endPage=6, ext={EN=ArticleExt(id=1322923595311219053, articleId=1322923593239232874, tenantId=1146029695717560320, journalId=1255847919539208197, language=EN, title=Principles of Processing Method Reducing Hepatotoxicity of Chuanlianzi（Toosendan Fructus）：Bioinformatics Prediction and Experiment Validation, columnId=1256183423710970242, journalTitle=Chinese Archives of Traditional Chinese Medicine, columnName=New Opinion of Ph.D. Supervisor, runingTitle=null, highlight=null, articleAbstract=
Objective

Based on network pharmacology and molecular docking techniques to predict the material basis and toxicity mechanism of Chuanlianzi（Toosendan Fructus）hepatotoxicity and to validate the reducing effects after processing in combination with in vivo and in vitro experiments.

Methods

The toxic components and targets of Chuanlianzi（Toosendan Fructus）were collected using the systematic pharmacology analysis platform of traditional Chinese medicine（TCMSP）and the Swiss Target Prediction database，respectively. Liver injury related targets were obtained through GeneCards and OMIM databases. After the common targets were screened by Venny diagram，PPI analysis was carried out by STRING database，and enrichment analysis on GO function and KEGG pathway were conducted to predict their possible toxicity mechanisms. Molecular docking validation and visualization of key components of Chuanlianzi（Toosendan Fructus）hepatotoxicity with key protein binding sites and binding effects were performed using AutoDockTools and PyMOL software. In vivo experiments were conducted to compare the magnitude of acute hepatotoxicity of different processed products of Chuanlianzi（Toosendan Fructus）in mice by detecting the viability of alanine aminotransferase（ALT）and aspartate aminotransferase（AST）in serum and observing the pathologic morphology of liver tissues with HE staining method and to determine the levels of superoxide dismutase（SOD），malondialdehyde（MDA），glutathione（GSH），and glutathione peroxidase（GSH-PX）and analyzed the reasons for the differences in the hepatotoxicity of different processed products of Chuanlianzi（Toosendan Fructus）in mice from the perspective of oxidative damage.

Results

Network pharmacology showed that Chuanlianzi（Toosendan Fructus）hepatotoxicity components were melianone，Toosendanin and Nimbolin A，and its core targets for liver injury were TP53，IL-6 and CASP3. GO enrichment was analyzed to the biological processes，such as positive regulation of transcription and gene expression，negative regulation of the apoptotic process，and inflammatory response，cellular components，such as extracellular region，outer mitochondrial membrane，mitochondria，protein binding，protein homodimerization activity，enzyme binding and other molecular functions. KEGG pathway analysis screened TNF，PI3K-Akt，IL-17，HIF-1，apoptosis and other key signaling pathways of Chuanlianzi（Toosendan Fructus）-induced hepatotoxicity. The molecular docking results showed that the main toxic components of Chuanlianzi（Toosendan Fructus）could bind well to the core targets. In vivo experimental results showed that compared with the raw Chuanlianzi（Toosendan Fructus），the vinegar-processing，stir-frying to brown and salt-processing products all significantly reduced the activities of ALT and AST in the serum of mice（P<0.05），and the wine-processing product showed a significant increase in both ALT and AST. HE staining showed that the vinegar-processing product，stir-frying to brown and salt-processing products reduced the infiltration of inflammatory cells in the liver tissues and apoptosis of hepatocytes of mice，which suggested that Chuanlianzi（Toosendan Fructus）had the effect of alleviating liver injury after processing. This suggested that Chuanlianzi（Toosendan Fructus）had the effect of reducing liver injury，but the hepatotoxicity of the wine-processing product was greater than that of the raw product. This result suggested that the mechanism of liver toxicity induced by raw Chuanlianzi（Toosendan Fructus）in mice was related to the disruption of their antioxidant defense system，reduced ability to scavenge free radicals and subsequent excessive accumulation of free radicals in the body，which induced cell membrane lipid peroxidation.

Conclusion

The vinegar-processing，stir-frying to brown and salt-processing products may reduce hepatotoxicity by altering the effects of Chuanlianzi（Toosendan Fructus）on hepatotoxic targets such as TP53，IL-6 and CASP3，reducing inflammatory responses and inhibition of cell membrane lipid peroxidation，but the wine-processing product is more toxic and should be avoided in clinical practice.

, authors=Chunhui ZENG, Lirong XING, Haipeng CHEN, Qunfeng LIN, Yu ZHANG, Hong CHEN, Ke YANG, authorsList=Chunhui ZENG, Lirong XING, Haipeng CHEN, Qunfeng LIN, Yu ZHANG, Hong CHEN, Ke YANG, authorCompany=null, correspAuthors=Ke YANG, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1322923609563464096, articleId=1322923593239232874, tenantId=1146029695717560320, journalId=1255847919539208197, language=CN, title=基于生物信息学预测川楝子肝毒性及实验探讨炮制减毒原理, columnId=1256183426563096985, journalTitle=中华中医药学刊, columnName=博士导师新论, runingTitle=null, highlight=null, articleAbstract=
目的

基于网络药理学和分子对接技术预测川楝子肝毒性的物质基础及毒性机制，并结合体内、外实验验证炮制减毒作用。

方法

利用中药系统药理学数据库与分析平台（TCMSP）与小分子药物预测作用靶点的在线平台（Swiss Target Prediction）分别收集川楝子的毒性成分和作用靶点。检索GeneCards和OMIM数据库获取肝损伤疾病靶点，绘制Venny图获取川楝子毒性成分和肝相关疾病的共同靶标后，利用STRING数据库进行蛋白-蛋白相互作用（PPI）网络分析，并对GO功能和KEGG通路进行富集分析，预测其可能的毒性机制。使用AutoDockTools和PyMOL软件对川楝子肝毒性的关键成分与关键蛋白结合位点及结合作用进行分子对接验证及可视化。体内实验通过检测小鼠血清中谷丙转氨酶（ALT）、谷草转氨酶（AST）的活力，苏木精-伊红染色法（HE）观察肝组织病理形态，比较川楝子不同炮制品致小鼠急性肝毒性的大小；测定小鼠肝脏中超氧化物歧物酶（SOD）、丙二醛（MDA）、还原型谷胱甘肽（GSH）、谷胱甘肽过氧化物酶（GSH-PX）的水平，从氧化损伤的角度分析川楝子不同炮制品致小鼠肝毒性大小差异的原因。

结果

网络药理学表明川楝子肝毒性成分为苦楝子酮、苦楝素、印楝波灵A等，其肝损伤核心靶点为肿瘤蛋白p53（TP53）、白细胞介素-6（IL-6）、半胱天冬蛋白酶-3（CASP3）等；GO富集分析到转录和基因表达的正调控、细胞凋亡过程的负调控、炎症反应等生物过程，胞外区、线粒体外膜、线粒体等细胞组分，蛋白质结合、蛋白质同源二聚活性、酶结合等分子功能；KEGG通路分析筛选到的肿瘤坏死因子（TNF）、磷脂酰肌醇3激酶（PI3K-Akt）、白细胞介素-17（IL-17）、缺氧诱导因子-1（HIF-1）、细胞凋亡等是川楝子引起肝毒性的关键信号通路。分子对接结果表明，川楝子主要毒性成分与核心靶点能较好结合。体内实验结果显示，与川楝子生品相比，醋炙品、炒焦品、盐炙品均能显著降低小鼠血清中ALT和AST的活性（P<0.05），酒炙品的ALT、AST均显著升高；HE染色结果显示，醋炙品、炒焦品、盐炙品可减少小鼠肝组织炎症细胞浸润及肝细胞的凋亡，提示川楝子炮制后具有减轻肝损伤的作用，但酒炙品的肝毒性要大于生品。此外，醋炙品、炒焦品、盐炙品可以降低生品造成的脂质过氧化，从而达到炮制减毒的效果，酒炙品则相反。

结论

醋炙、炒焦、盐炙炮制方法可能通过改变川楝子对TP53、IL-6和CASP3等肝毒性靶点的影响，减轻炎症反应，抑制细胞膜脂质过氧化，从而降低肝毒性，但酒炙方法毒性增强，在临床上应该避免。

, authors=曾春晖, 邢丽蓉, 陈海鹏, 林群峰, 章昱, 陈泓, 杨柯, authorsList=曾春晖, 邢丽蓉, 陈海鹏, 林群峰, 章昱, 陈泓, 杨柯, authorCompany=null, correspAuthors=杨柯, authorNote=

曾春晖（1974-），女，广西钦州人，教授，博士研究生导师，博士，研究方向：中药炮制原理。

, correspAuthorsNote=
杨柯（1975-），男（侗族），广西柳州人，教授，硕士研究生导师，硕士，研究方向：中药抗炎免疫药理。E-mail：。
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曾春晖（1974-），女，广西钦州人，教授，博士研究生导师，博士，研究方向：中药炮制原理。

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曾春晖（1974-），女，广西钦州人，教授，博士研究生导师，博士，研究方向：中药炮制原理。

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experiment［J］．Chin J Mod Appl Pharm，2022，39（16）：2052-2062., articleTitle=Study on the toxicological mechanism of psoraleaefructus induced liver injury based on network pharmacology and cell experiment, refAbstract=null), Reference(id=1322923675871216189, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, doi=null, pmid=null, pmcid=null, year=2023, volume=15, issue=377, pageStart=51, pageEnd=61, url=null, language=null, rfNumber=[25], rfOrder=24, authorNames=LIANG Y, CHEN S, HAN S, journalName=Toxicol Lett, refType=null, unstructuredReference=LIANG Y，CHEN S，HAN S，et al．Toosendanin induced hepatotoxicity via triggering PERK-eIF2α-ATF4 mediated ferroptosis［J］．Toxicol Lett，2023，15（377）：51-61., articleTitle=Toosendanin induced hepatotoxicity via triggering PERK-eIF2α-ATF4 mediated ferroptosis, refAbstract=null), Reference(id=1322923676315812415, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, doi=null, pmid=null, pmcid=null, year=2020, volume=null, issue=null, pageStart=null, pageEnd=null, url=null, language=null, rfNumber=[26], rfOrder=25, authorNames=王蔚倩, journalName=null, refType=null, unstructuredReference=王蔚倩．菊三七及主要毒性成分千里光碱致肝窦内皮损伤的毒性机制探讨［D］．上海：上海中医药大学，2020., articleTitle=菊三七及主要毒性成分千里光碱致肝窦内皮损伤的毒性机制探讨, refAbstract=null), Reference(id=1322923677041427008, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, doi=null, pmid=null, pmcid=null, year=2022, volume=2022, issue=null, pageStart=null, pageEnd=null, url=null, language=null, rfNumber=[27], rfOrder=26, authorNames=XING B, CUI Y, CHEN Y, journalName=Evid Based Complement Alternat Med, refType=null, unstructuredReference=XING B，CUI Y，CHEN Y，et al．Exploration of the components and pharmacological mechanisms of keyin pill-induced liver injury based on network pharmacology［J］．Evid Based Complement Alternat Med，2022，2022：9916949., articleTitle=Exploration of the components and pharmacological 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序号成分中文名称分子式MOL IDOB/（%）OL
CLZ1Mandenol甘露醇C20H36O2MOL00149442.000.19
CLZ2Ethyl linolenate亚麻酸乙酯C20H34O2MOL00149546.100.20
CLZ3Stigmasterol豆甾醇C29H48OMOL00204543.410.76
CLZ4Melianone苦楝子酮C30H46O4MOL00204740.730.81
CLZ5Nimbolidin D印楝波为定DC44H58O12MOL00204830.380.53
CLZ6Nimbolin A印楝波灵AC39H46O8MOL00205332.110.34
CLZ7（E）-3-［（2S，3R）-2-（4-hydroxy-3-methoxy-phenyl）-7-methoxy-3-methylol-2，3-dihydrobenzofuran-5-yl］acrolein—C20H20O6MOL00205654.740.40
CLZ8Medioresil皮树脂醇C21H24O7MOL00205857.200.62
CLZ9Quercetin槲皮素C15H10O7MOL00009846.430.28
CLZ10Nimbolidin B印楝波为定BC38H50O12———
CLZ11Nimbolidin C印楝波为定CC37H50O12———
CLZ12Nimbolidin E印楝波为定EC43H58O12———
CLZ13Toosendanin苦楝素C30H38O11———
), ArticleFig(id=1322923645286351367, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=CN, label=表1, caption=

TCMSP数据库筛选的川楝子主要活性成分

, figureFileSmall=null, figureFileBig=null, tableContent=
序号成分中文名称分子式MOL IDOB/（%）OL
CLZ1Mandenol甘露醇C20H36O2MOL00149442.000.19
CLZ2Ethyl linolenate亚麻酸乙酯C20H34O2MOL00149546.100.20
CLZ3Stigmasterol豆甾醇C29H48OMOL00204543.410.76
CLZ4Melianone苦楝子酮C30H46O4MOL00204740.730.81
CLZ5Nimbolidin D印楝波为定DC44H58O12MOL00204830.380.53
CLZ6Nimbolin A印楝波灵AC39H46O8MOL00205332.110.34
CLZ7（E）-3-［（2S，3R）-2-（4-hydroxy-3-methoxy-phenyl）-7-methoxy-3-methylol-2，3-dihydrobenzofuran-5-yl］acrolein—C20H20O6MOL00205654.740.40
CLZ8Medioresil皮树脂醇C21H24O7MOL00205857.200.62
CLZ9Quercetin槲皮素C15H10O7MOL00009846.430.28
CLZ10Nimbolidin B印楝波为定BC38H50O12———
CLZ11Nimbolidin C印楝波为定CC37H50O12———
CLZ12Nimbolidin E印楝波为定EC43H58O12———
CLZ13Toosendanin苦楝素C30H38O11———
), ArticleFig(id=1322923645999383050, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
有效活性成分核心靶点结合自由能（ka/mol）
MelianoneTP53-6.19
IL-6-4.82
CASP3-4.58
ESR1-5.79
HIF1A-5.83
Nimbolin ATP53-3.49
IL-6-2.63
CASP3-4.71
ESR1-4.61
HIF1A-4.67
ToosendaninTP53-2.75
IL-6-2.73
CASP3-3.17
ESR1-5.10
HIF1A-4.01
), ArticleFig(id=1322923646439784972, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=CN, label=表2, caption=

川楝子核心成分分子对接能量

, figureFileSmall=null, figureFileBig=null, tableContent=
有效活性成分核心靶点结合自由能（ka/mol）
MelianoneTP53-6.19
IL-6-4.82
CASP3-4.58
ESR1-5.79
HIF1A-5.83
Nimbolin ATP53-3.49
IL-6-2.63
CASP3-4.71
ESR1-4.61
HIF1A-4.67
ToosendaninTP53-2.75
IL-6-2.73
CASP3-3.17
ESR1-5.10
HIF1A-4.01
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组别n剂量/（g·kg-1）ALT/（IU/L）AST/（IU/L）
正常组10—10.80±2.2321.55±5.22
生品组10MTD24.72±1.75**46.80±6.41**
醋炙品组10MTD19.26±1.81**##46.44±9.29**
酒炙品组10MTD38.76±2.69**77.11±23.43**
炒焦品组10MTD21.20±3.22**#48.53±5.34**
盐炙品组10MTD21.36±3.39*34.66±2.53**##
), ArticleFig(id=1322923648591462927, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=CN, label=表3, caption=

川楝子不同炮制品给药后2 h对小鼠ALT和AST活力的影响（±s）

, figureFileSmall=null, figureFileBig=null, tableContent=
组别n剂量/（g·kg-1）ALT/（IU/L）AST/（IU/L）
正常组10—10.80±2.2321.55±5.22
生品组10MTD24.72±1.75**46.80±6.41**
醋炙品组10MTD19.26±1.81**##46.44±9.29**
酒炙品组10MTD38.76±2.69**77.11±23.43**
炒焦品组10MTD21.20±3.22**#48.53±5.34**
盐炙品组10MTD21.36±3.39*34.66±2.53**##
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组别n剂量/（g·kg-1）SOD/（U/mgprot）GSH-Px/（U/mgprot）CAT/（U/mgprot）XOD/（U/mgprot）
正常组10—355.8±54.8586.2±111.489.24±18.388.80±0.75
生品组10MTD378.2±26.2450.4±93.0*37.98±10.77**12.38±0.84**
100.852MTD343.3±56.7608.1±162.556.68±12.91**12.23±1.44*
100.854MTD382.6±21.4478.8±97.689.45±15.9110.43±0.91**
醋炙品组10MTD414.5±59.6688.3±93.046.95±9.00**9.54±1.10
100.85MTD368.5±88.4573.8±236.341.13±7.03**10.39±2.06
100.852MTD363.1±85.5526.4±161.470.27±16.8710.28±2.00
酒炙品组10MTD367.4±32.8455.2±85.0*55.66±19.28**10.39±0.82**
100.852MTD435.3±111.6447.2±56.4**61.86±15.38*10.46±1.68
100.855MTD495.8±84.1458.1±67.372.91±36.229.36±1.30
炒焦品组10MTD353.0±42.5501.2±91.544.42±12.85**10.05±0.88*
100.85MTD430.0±87.1433.1±117.1*49.45±13.899.10±1.86
100.852MTD326.1±37.7644.0±137.866.91±24.739.71±0.96
盐炙品组10MTD398.7±64.4591.7±121.146.40±16.15**9.12±1.14
100.85MTD359.1±104.2559.1±126.472.40±15.599.97±2.52
100.852MTD383.3±84.0622.7±189.6115.13±37.9511.15±3.45
), ArticleFig(id=1322923649946223122, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=CN, label=表4, caption=

川楝子不同炮制品对小鼠肝组织氧化应激相关酶活性的影响（±s）

, figureFileSmall=null, figureFileBig=null, tableContent=
组别n剂量/（g·kg-1）SOD/（U/mgprot）GSH-Px/（U/mgprot）CAT/（U/mgprot）XOD/（U/mgprot）
正常组10—355.8±54.8586.2±111.489.24±18.388.80±0.75
生品组10MTD378.2±26.2450.4±93.0*37.98±10.77**12.38±0.84**
100.852MTD343.3±56.7608.1±162.556.68±12.91**12.23±1.44*
100.854MTD382.6±21.4478.8±97.689.45±15.9110.43±0.91**
醋炙品组10MTD414.5±59.6688.3±93.046.95±9.00**9.54±1.10
100.85MTD368.5±88.4573.8±236.341.13±7.03**10.39±2.06
100.852MTD363.1±85.5526.4±161.470.27±16.8710.28±2.00
酒炙品组10MTD367.4±32.8455.2±85.0*55.66±19.28**10.39±0.82**
100.852MTD435.3±111.6447.2±56.4**61.86±15.38*10.46±1.68
100.855MTD495.8±84.1458.1±67.372.91±36.229.36±1.30
炒焦品组10MTD353.0±42.5501.2±91.544.42±12.85**10.05±0.88*
100.85MTD430.0±87.1433.1±117.1*49.45±13.899.10±1.86
100.852MTD326.1±37.7644.0±137.866.91±24.739.71±0.96
盐炙品组10MTD398.7±64.4591.7±121.146.40±16.15**9.12±1.14
100.85MTD359.1±104.2559.1±126.472.40±15.599.97±2.52
100.852MTD383.3±84.0622.7±189.6115.13±37.9511.15±3.45
), ArticleFig(id=1322923650659254804, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=EN, label=null, caption=null, figureFileSmall=null, figureFileBig=null, tableContent=
组别n剂量/（g·kg-1）GSH/（μmol/L）MDA/（nmol/mgprot）
正常组10—35.93±13.120.490±0.095
生品组10MTD15.21±2.86**0.909±0.171**
100.852MTD26.77±9.940.494±0.086
100.854MTD33.70±13.340.613±0.144
醋炙品组10MTD24.52±9.230.687±0.187*
100.85MTD26.64±14.670.505±0.133
100.852MTD23.62±6.230.515±0.099
酒炙品组10MTD26.88±5.910.789±0.215*
100.852MTD21.17±8.280.590±0.232
100.855MTD22.09±15.240.564±0.102
炒焦品组10MTD19.46±8.01*0.674±0.237
100.85MTD36.11±8.810.697±0.311
100.852MTD38.27±22.880.607±0.135
盐炙品组10MTD37.71±16.520.743±0.246
100.85MTD40.41±17.400.668±0.226
100.852MTD48.73±18.410.660±0.205
), ArticleFig(id=1322923651514892821, tenantId=1146029695717560320, journalId=1255847919539208197, articleId=1322923593239232874, language=CN, label=表5, caption=

川楝子不同炮制品对小鼠肝组织氧化应激产物含量的影响（±s）

, figureFileSmall=null, figureFileBig=null, tableContent=
组别n剂量/（g·kg-1）GSH/（μmol/L）MDA/（nmol/mgprot）
正常组10—35.93±13.120.490±0.095
生品组10MTD15.21±2.86**0.909±0.171**
100.852MTD26.77±9.940.494±0.086
100.854MTD33.70±13.340.613±0.144
醋炙品组10MTD24.52±9.230.687±0.187*
100.85MTD26.64±14.670.505±0.133
100.852MTD23.62±6.230.515±0.099
酒炙品组10MTD26.88±5.910.789±0.215*
100.852MTD21.17±8.280.590±0.232
100.855MTD22.09±15.240.564±0.102
炒焦品组10MTD19.46±8.01*0.674±0.237
100.85MTD36.11±8.810.697±0.311
100.852MTD38.27±22.880.607±0.135
盐炙品组10MTD37.71±16.520.743±0.246
100.85MTD40.41±17.400.668±0.226
100.852MTD48.73±18.410.660±0.205
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基于生物信息学预测川楝子肝毒性及实验探讨炮制减毒原理
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曾春晖 , 邢丽蓉 , 陈海鹏 , 林群峰 , 章昱 , 陈泓 , 杨柯
中华中医药学刊 | 博士导师新论 2025,43(4): 1-6
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中华中医药学刊 |博士导师新论 2025 , 43 (4) : 1 -6
基于生物信息学预测川楝子肝毒性及实验探讨炮制减毒原理
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曾春晖（1974-），女，广西钦州人，教授，博士研究生导师，博士，研究方向：中药炮制原理。

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曾春晖（1974-），女，广西钦州人，教授，博士研究生导师，博士，研究方向：中药炮制原理。

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曾春晖, 邢丽蓉, 陈海鹏, 林群峰, 章昱, 陈泓, 杨柯
作者信息
  • 广西中医药大学药学院，广西　南宁　530200
通讯作者:
杨柯（1975-），男（侗族），广西柳州人，教授，硕士研究生导师，硕士，研究方向：中药抗炎免疫药理。E-mail：。
作者简介:

曾春晖（1974-），女，广西钦州人，教授，博士研究生导师，博士，研究方向：中药炮制原理。

Principles of Processing Method Reducing Hepatotoxicity of Chuanlianzi（Toosendan Fructus）：Bioinformatics Prediction and Experiment Validation
Chunhui ZENG, Lirong XING, Haipeng CHEN, Qunfeng LIN, Yu ZHANG, Hong CHEN, Ke YANG
Affiliations
  • College of Pharmacy, Guangxi University of Chinese Medicine, Nanning 530200, Guangxi, China
出版时间: 2025-04-10 doi: 10.13193/j.issn.1673-7717.2025.04.001
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摘要
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目的

基于网络药理学和分子对接技术预测川楝子肝毒性的物质基础及毒性机制，并结合体内、外实验验证炮制减毒作用。

方法

利用中药系统药理学数据库与分析平台（TCMSP）与小分子药物预测作用靶点的在线平台（Swiss Target Prediction）分别收集川楝子的毒性成分和作用靶点。检索GeneCards和OMIM数据库获取肝损伤疾病靶点，绘制Venny图获取川楝子毒性成分和肝相关疾病的共同靶标后，利用STRING数据库进行蛋白-蛋白相互作用（PPI）网络分析，并对GO功能和KEGG通路进行富集分析，预测其可能的毒性机制。使用AutoDockTools和PyMOL软件对川楝子肝毒性的关键成分与关键蛋白结合位点及结合作用进行分子对接验证及可视化。体内实验通过检测小鼠血清中谷丙转氨酶（ALT）、谷草转氨酶（AST）的活力，苏木精-伊红染色法（HE）观察肝组织病理形态，比较川楝子不同炮制品致小鼠急性肝毒性的大小；测定小鼠肝脏中超氧化物歧物酶（SOD）、丙二醛（MDA）、还原型谷胱甘肽（GSH）、谷胱甘肽过氧化物酶（GSH-PX）的水平，从氧化损伤的角度分析川楝子不同炮制品致小鼠肝毒性大小差异的原因。

结果

网络药理学表明川楝子肝毒性成分为苦楝子酮、苦楝素、印楝波灵A等，其肝损伤核心靶点为肿瘤蛋白p53（TP53）、白细胞介素-6（IL-6）、半胱天冬蛋白酶-3（CASP3）等；GO富集分析到转录和基因表达的正调控、细胞凋亡过程的负调控、炎症反应等生物过程，胞外区、线粒体外膜、线粒体等细胞组分，蛋白质结合、蛋白质同源二聚活性、酶结合等分子功能；KEGG通路分析筛选到的肿瘤坏死因子（TNF）、磷脂酰肌醇3激酶（PI3K-Akt）、白细胞介素-17（IL-17）、缺氧诱导因子-1（HIF-1）、细胞凋亡等是川楝子引起肝毒性的关键信号通路。分子对接结果表明，川楝子主要毒性成分与核心靶点能较好结合。体内实验结果显示，与川楝子生品相比，醋炙品、炒焦品、盐炙品均能显著降低小鼠血清中ALT和AST的活性（P<0.05），酒炙品的ALT、AST均显著升高；HE染色结果显示，醋炙品、炒焦品、盐炙品可减少小鼠肝组织炎症细胞浸润及肝细胞的凋亡，提示川楝子炮制后具有减轻肝损伤的作用，但酒炙品的肝毒性要大于生品。此外，醋炙品、炒焦品、盐炙品可以降低生品造成的脂质过氧化，从而达到炮制减毒的效果，酒炙品则相反。

结论

醋炙、炒焦、盐炙炮制方法可能通过改变川楝子对TP53、IL-6和CASP3等肝毒性靶点的影响，减轻炎症反应，抑制细胞膜脂质过氧化，从而降低肝毒性，但酒炙方法毒性增强，在临床上应该避免。

川楝子  /  生物信息学  /  肝毒性  /  炮制减毒  /  实验验证
Objective

Based on network pharmacology and molecular docking techniques to predict the material basis and toxicity mechanism of Chuanlianzi（Toosendan Fructus）hepatotoxicity and to validate the reducing effects after processing in combination with in vivo and in vitro experiments.

Methods

The toxic components and targets of Chuanlianzi（Toosendan Fructus）were collected using the systematic pharmacology analysis platform of traditional Chinese medicine（TCMSP）and the Swiss Target Prediction database，respectively. Liver injury related targets were obtained through GeneCards and OMIM databases. After the common targets were screened by Venny diagram，PPI analysis was carried out by STRING database，and enrichment analysis on GO function and KEGG pathway were conducted to predict their possible toxicity mechanisms. Molecular docking validation and visualization of key components of Chuanlianzi（Toosendan Fructus）hepatotoxicity with key protein binding sites and binding effects were performed using AutoDockTools and PyMOL software. In vivo experiments were conducted to compare the magnitude of acute hepatotoxicity of different processed products of Chuanlianzi（Toosendan Fructus）in mice by detecting the viability of alanine aminotransferase（ALT）and aspartate aminotransferase（AST）in serum and observing the pathologic morphology of liver tissues with HE staining method and to determine the levels of superoxide dismutase（SOD），malondialdehyde（MDA），glutathione（GSH），and glutathione peroxidase（GSH-PX）and analyzed the reasons for the differences in the hepatotoxicity of different processed products of Chuanlianzi（Toosendan Fructus）in mice from the perspective of oxidative damage.

Results

Network pharmacology showed that Chuanlianzi（Toosendan Fructus）hepatotoxicity components were melianone，Toosendanin and Nimbolin A，and its core targets for liver injury were TP53，IL-6 and CASP3. GO enrichment was analyzed to the biological processes，such as positive regulation of transcription and gene expression，negative regulation of the apoptotic process，and inflammatory response，cellular components，such as extracellular region，outer mitochondrial membrane，mitochondria，protein binding，protein homodimerization activity，enzyme binding and other molecular functions. KEGG pathway analysis screened TNF，PI3K-Akt，IL-17，HIF-1，apoptosis and other key signaling pathways of Chuanlianzi（Toosendan Fructus）-induced hepatotoxicity. The molecular docking results showed that the main toxic components of Chuanlianzi（Toosendan Fructus）could bind well to the core targets. In vivo experimental results showed that compared with the raw Chuanlianzi（Toosendan Fructus），the vinegar-processing，stir-frying to brown and salt-processing products all significantly reduced the activities of ALT and AST in the serum of mice（P<0.05），and the wine-processing product showed a significant increase in both ALT and AST. HE staining showed that the vinegar-processing product，stir-frying to brown and salt-processing products reduced the infiltration of inflammatory cells in the liver tissues and apoptosis of hepatocytes of mice，which suggested that Chuanlianzi（Toosendan Fructus）had the effect of alleviating liver injury after processing. This suggested that Chuanlianzi（Toosendan Fructus）had the effect of reducing liver injury，but the hepatotoxicity of the wine-processing product was greater than that of the raw product. This result suggested that the mechanism of liver toxicity induced by raw Chuanlianzi（Toosendan Fructus）in mice was related to the disruption of their antioxidant defense system，reduced ability to scavenge free radicals and subsequent excessive accumulation of free radicals in the body，which induced cell membrane lipid peroxidation.

Conclusion

The vinegar-processing，stir-frying to brown and salt-processing products may reduce hepatotoxicity by altering the effects of Chuanlianzi（Toosendan Fructus）on hepatotoxic targets such as TP53，IL-6 and CASP3，reducing inflammatory responses and inhibition of cell membrane lipid peroxidation，but the wine-processing product is more toxic and should be avoided in clinical practice.

Chuanlianzi（Toosendan Fructus）  /  bioinformatics  /  hepatotoxicity  /  reducing toxin after processing  /  experiment validation
曾春晖, 邢丽蓉, 陈海鹏, 林群峰, 章昱, 陈泓, 杨柯. 基于生物信息学预测川楝子肝毒性及实验探讨炮制减毒原理. 中华中医药学刊, 2025 , 43 (4) : 1 -6 . DOI: 10.13193/j.issn.1673-7717.2025.04.001
Chunhui ZENG, Lirong XING, Haipeng CHEN, Qunfeng LIN, Yu ZHANG, Hong CHEN, Ke YANG. Principles of Processing Method Reducing Hepatotoxicity of Chuanlianzi（Toosendan Fructus）：Bioinformatics Prediction and Experiment Validation[J]. Chinese Archives of Traditional Chinese Medicine, 2025 , 43 (4) : 1 -6 . DOI: 10.13193/j.issn.1673-7717.2025.04.001
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川楝子为楝科植物川楝Melia toosendan Sieb. et Zucc.的干燥成熟果实，味苦，寒，有小毒，归肝、小肠、膀胱经，有疏肝泄热，行气止痛，杀虫之功效[1]，用于肝郁化火，胸肋、脘腹胀痛，疝气疼痛，虫积腹痛。川楝子始载于《神农本草经》，列于下品，是理气和传统的驱虫药。2020版《中华人民共和国药典》收录了生川楝子和炒川楝子2个炮制品[1]。毒理学研究发现，虽然川楝子作用众多，但其提取物可以诱导急性肝损伤[2-3]。有资料显示，民间使用川楝子进行驱虫时，发现有28例发生肝中毒现象，10例死亡病例[4]，限制了川楝子的使用。目前，基于炮制减毒研究川楝子肝损伤的报道较少，因此探讨不同炮制品的炮制减毒原理具有重要意义。网络药理学是基于分析生物网络设计药物分子的一门学科，通过系统生物学、生物信息学、多组学联用等多学科的理念，采用高通量组学数据分析、计算机虚拟计算及网络数据库检索等方式以实现化合物-靶点-信号通路可视化，可在一定程度上揭示天然植物中各活性成分作用功效[5]，是建立在药物-疾病-靶点多层次的网络基础上的新兴领域，主要从多种靶点的相互作用来研究疾病的治疗，广泛用于医学领域[6]。川楝子的化学成分众多且复杂，呈现多成分、多靶点相互协同作用，需综合分析其药理活性及作用机制。为探讨川楝子引起肝毒性的机制及炮制减毒原理，本研究采用网络药理学的研究方法，将川楝子的肝毒性成分可能作用靶点与肝损伤机制相关联，预测川楝子致肝毒性的靶点和通路，将川楝子的肝毒性成分和核心靶点进行分子对接分析，同时通过体内、外实验比较川楝子不同炮制品肝毒性大小，分析炮制减毒原理，为临床安全使用提供实验依据。
1　材料与方法
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1.1　川楝子主要成分的筛选及作用靶点预测
采用中药系统药理学数据库与分析平台（TCMSP）[7]（https://tcmspw.com/tcmsp），以“川楝子”为关键词搜索，同时满足口服生物利用度（OB）≥30%，类药性（DL）≥0.18筛选川楝子的主要成分。在TCMSP平台中，收集已检索成分的靶点，并通过SwissTargetPrediction平台（http://www.swisstargetprediction.ch/），检索补充川楝子的主要成分靶点，进行整合及去除重复项，所得靶点与Uniprot数据库（https://www.uniprot.org/）进行匹配，得到各个靶点的基因名称。
1.2　疾病作用靶点预测
以“Liver toxic”和“Liver toxicity”为关键词在OMIM数据库[8]（https://omim.org/）、GeneCards数据库（https://www.genecards.org/）中搜索、整合肝损伤疾病靶点，并用Venny图取川楝子靶点和肝毒性靶点的交集。
1.3　川楝子药物成分-疾病-靶点网络构建
网络拓扑数据分析软件Cytoscape[9]是一款图形化显示网络并进行分析和编辑的软件，可以利用自身以及第三方开发的大量功能插件针对网络问题进行深入分析。将“1.1”和“1.2”项得到的数据导入Cytoscape软件中，构建川楝子药物-疾病-靶点网络图。
1.4　PPI网络的构建与核心靶点的筛选
为明确川楝子各靶点间相互作用，将其潜在作用靶点导入STRING数据库[10]（http:l/lstring-db.org/），设置Organism为“homo sapines”后进行搜索，构建蛋白-蛋白相互作用（PPI）网络，根据靶点连线的密集程度判断靶点之间的相互作用程度；将其导入Cytoscape软件进行可视化分析，根据度值（degree）大小进一步设置节点大小与颜色，度值大小与节点大小和颜色深浅成正比，得到PPI网络。
1.5　GO富集分析和KEGG通路富集分析
利用DAVID数据库对差异基因进行GO和KEGG途径富集分析[11]。GO富集分析包括细胞成分（CC）分析、生物过程（BP）分析和分子功能（MF）分析。检索时输入差异基因，标识符选择“OFFICIAL_GENE_SYMBOL”，物种选择“Homo sapiens”，并对分析结果进行可视化处理。
1.6　分子对接
1.6.1　配体的准备
经过1.1中的核心成分筛选，收集到已被证实的川楝子肝毒性化学成分，得到川楝子肝毒性成分列表，见表1。基于Pubchem数据库[12]（https://pubchem.ncbi.nlm.nih.gov/）下载川楝子化学结构的sdf.格式文件，并通过Open Bable 2.4.1软件转化为pdb.格式文件，采用AutoDock-Tools软件设置其可旋转的键及活动键时需要移动的最少的原子数目，输出pdbqt.格式文件作为分子对接中的配体[13]。
1.6.2　受体的准备
首先对其结构进行预处理，去除水分子及配体，然后采用AutoDockTools软件计算Gasteiger电荷，添加氢原子，分离配体和受体等操作，输出pdbqt.格式作为分子对接中的受体。
1.6.3　分子对接
将受体与配体导入AutoDockTools软件中获取相应毒性口袋，设置Gridbox坐标及大小，运行AutoDock进行分子对接，计算药物活性成分与靶蛋白结合所需的最低结合能。结果以化合物分子与受体结合能量是否为负值进行结合的评价标准，负值时表明两者可以结合，且能量越低，构象越稳定，发生的作用可能性则越大，其对接结果越可靠。最后，通过开源软件PyMOL对其对接结果进行可视化。
1.7　实验验证
1.7.1　动物和细胞
SPF级健康昆明种小鼠，雌雄各半，体质量（20±2）g。小鼠自由摄食饮水，均购自广西医科大学实验动物中心，动物许可证号：SCXK桂20140002。人正常肝细胞株L-02，购买于上海美轩生物科技有限公司。
1.7.2　药材与试剂
川楝子药材购自玉林药市，经广西中医药大学药用植物教研室郭敏副教授鉴定为楝科植物川楝Melia toosendan Sieb. et Zucc.的干燥成熟果实，批号20161120。
丙氨酸转移酶（ALT）、门冬氨酸氨基转移酶（AST）：南京建成生物研究所20161206；超氧化物歧化酶（SOD）：南京建成生物研究所20170603；丙二醛（MDA）：南京建成生物研究所20170602；胱甘肽（GSH）：南京建成生物研究所20170601；谷胱甘肽过氧化物酶（GSH-Px）：南京建成生物研究所20170306；过氧化氢酶（CAT）：南京建成生物研究所20170605；黄嘌呤氧化酶（XOD）：南京建成生物研究所20170602；无水乙醇AR：成都科隆化学品有限公司2017071301。胎牛血清：浙江天坑生物科技股份有限公司20170223；RPMI-1640不完全培养基：江苏凯基生物技术股份有限公司20170926；胰蛋白酶-EDTA消化液：solarbio公司20170915；噻唑兰（MTT）：solarbio公司804W054；二甲基亚砜（DMSO）：sigma公司1029B038；PBS：solarbio公司20170728；线粒体复合物酶Ⅰ～Ⅳ：solarbio公司20180115。
1.7.3　炮制品的制备
生品：取川楝子，除去杂质，即得[1]。炒焦品：取生川楝子，切厚片或碾碎，置热锅中，用中火炒至表面焦黄色，断面色加深，焦斑为1/2～1/4为度，取出，放晾，本品呈半球状、厚片或不规则的碎块，表面焦黄色，气焦香，味酸、苦[14]。酒炙品：取生川楝子1 kg，加入约250 mL的高度白酒（含乙醇50%～60%）浸润，待白酒吸干后，用小火炒至色泽微黄[15]。醋炙品：取生川楝子1 kg，加15%醋拌润，待醋液吸尽后，文火炒干放冷[16]。盐炙品：取生川楝子碎块1 kg，加入20 g食盐用4～5倍量水稀释，稍闷，待盐水被吸尽后，置热锅内，用文火加热，炒至深黄色，取出，晾凉[17]。
1.7.4　供试品制备
取川楝子5种炮制品各约1 kg，加10倍量水浸泡3 h，加热煎煮2 h取出，用8层纱布过滤后，药渣内加入8倍量水，再煎煮0.5 h，取出用8层纱布过滤，合并煎煮液后浓缩转移至敞口容器中，加热蒸发至浸膏状，转移至干燥的蒸发皿中，水浴持续加热直至成为黏稠的难以流动的稠浸膏，生品得膏率为21.33%；醋炙品得膏率为22.85%；酒炙品得膏率为15.55%；炒焦品得膏率为19.71%；盐炙品得膏率为20.75%，于冰箱4℃中冷藏保存。
1.7.5　仪器
NM32LA型氮气发生器（英国Peak scientific公司）；BP211D型电子天平（德国Sartorius公司）；DETD-200型手提高速万能粉碎机（中国浙江温岭市林大机械有限公司）；BGZ-76型电热鼓风干燥器（上海白典仪器设备有限公司）。HR1200-ⅡB2型生物安全柜（青岛海尔特种电器有限公司）；IX2-SLP型倒置显微镜（日本OLYMPUS公司）；ST16R型冷冻低速离心机（美国Thermo公司）；HVA-110型高压灭菌锅（日本Hirayama公司）；MCO-18AIC型恒温二氧化碳培养箱（日本Panasonic松下公司）；KB-900型定轨摇床（江苏海门其林贝尔仪器制造公司）。
1.7.6　川楝子不同炮制品最大耐受量（MTD）测定
前期川楝子各炮制品的最大耐受量测定得川楝子生品MTD值为184 g·kg-1，醋炙品MTD值为179 g·kg-1，酒炙品MTD值为247 g·kg-1，炒焦品MTD值为136 g·kg-1，盐炙品MTD值为198 g·kg-1。取KM小鼠60只，雌雄各半，每组10只，分为正常组、生品组、醋炙品组、酒炙品组、盐炙品组、炒焦品组，禁食不禁水12 h，每只小鼠灌胃给药40 mL·kg-1。正常组小鼠灌胃给予等体积蒸馏水。一日内给药3次，间隔6 h，给药后2 h，称重，眼眶静脉丛取血，3000 r/min，离心10 min，取血清，测定血清ALT、AST活力，肝组织经苏木精-伊红染色法（HE）染色，镜下观察其组织形态学变化。
1.7.7　致小鼠肝毒性氧化损伤的实验研究
根据前期研究结果[18]，采用各炮制品的最大中毒剂量、亚中毒剂量、最大无毒剂量作为肝毒性氧化损伤实验的高中低剂量的选择。取KM小鼠160只，雌雄各半，每组10只，分为正常组、生品组、醋炙品组、酒炙品组、盐炙品组、炒焦品组，禁食不禁水12 h，每只小鼠灌胃给药40 mL·kg-1，间隔6 h给药3次。正常组小鼠灌予同样体积的蒸馏水。生品组设置3个剂量组，分别为MTD、0.852MTD、0.854MTD；醋炙品组、炒焦品组、盐炙品组设置3个剂量组，分别为MTD、0.85MTD、0.852MTD；酒炙品组设置3个剂量组，分别为MTD、0.852MTD、0.855MTD。给药后2 h，取小鼠肝组织，加冷生理盐水制成10%肝匀浆，离心取上清液，检测肝组织中SOD、CAT、XOD、GSH-Px活性及MDA、GSH含量。
1.7.8　数据处理
数据用表示，采用SPSS 17.0统计软件对各组参数进行单因素方差分析，进行组间比较，P<0.05为数据差异有统计学意义。
2　结果
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2.1　药物成分与靶点筛选
在TCMSP数据库中检索川楝子的活性成分，同时满足OB≥30%，DL≥0.18共筛选出川楝子活性成分9个，活性成分对应靶点275个。此外，还有4种成分并未收录到该数据库中，分别为Nimbolidin B、Nimbolidin C、Nimbolidin E、Toosendanin，共得到13种成分，见表1。在Swiss Target Prediction数据库中检索川楝子中这4种活性成分的靶点，最终得到活性成分对应靶点131个，将潜在靶点导入Uniprot数据库进行基因名称的校正，得到药物的主要作用靶点共406个，并纳入分析。
2.2　疾病靶点筛选
分别在GeneCards数据库、OMIM数据库中筛选并获得2473个肝毒性靶点，将川楝子的靶点与肝毒性靶点整理后，利用Venny图取交集，得川楝子致肝损伤有效靶点128个，如插页Ⅰ图1所示。
2.3　川楝子药物-成分-疾病-靶点网络构建结果
将“2.1”项得到的川楝子13个成分与128个交集靶点导入Cytoscape中，构建川楝子药物-成分-疾病靶点网络，并进行网络拓扑分析，如插页Ⅰ图2，该网络共有142个节点，141条边。NetworkAnalyzer功能分析显示该网络中度（degree）值排名前13的成分见表1。
2.4　PPI网络的构建
通过String数据库分析128个映射得到的交集靶点，将数据进行分析之后导入Cytoscape软件，构建预测川楝子肝毒性靶点PPI网络，如插页Ⅰ图3所示，图中颜色越深，面积越大，表示靶点度值越大，在网络中越重要。
2.5　GO与KEGG富集分析
GO功能富集分析结果显示，得到492条生物过程（biologicalprocess，BP），56条细胞组成（cellularcomponent，CC）和109条分子功能（molecularfunction，MF），将部分条目可视化，如插页Ⅱ图4。川楝子主要涉及基因表达的正调控、RNA聚合酶Ⅱ启动子的转录正调控、细胞凋亡过程的负调控、炎症反应等生物过程；胞外区、细胞外间隙、大分子复合物、线粒体外膜、核质、线粒体等细胞组成；蛋白结合、相同蛋白结合、RNA聚合酶Ⅱ序列特异性DNA结合转录因子结合等蛋白组成。KEGG通路富集分析显示，川楝子肝毒性的相关通路共133条，将排名前20的通路可视化，如插页Ⅱ图5，主要涉及癌症通路、乙型肝炎、TNF信号通路、非小细胞肺癌、糖尿病并发症中的AGERAGE信号通路、PI3K-Akt信号通路、IL-17信号通路、HIF-1信号通路、细胞凋亡、肝细胞癌等通路。
2.6　分子对接
选择能与肝损伤靶点结合且度值排名在前的3个关键成分，分别与TP53、IL-6、CASP3、ESR1、HIF1A的分子对接结合信息见表2。表中结合自由能表示受体和配体之间的结合活性，由表可知关键成分与核心靶点对接的结合能均为负值。使用PyMOL对部分对接结果作可视化示例，如插页Ⅱ图6。
2.7　实验结果
2.7.1　对血清ALT、AST的影响
由表3结果显示，与正常组相比，川楝子生品组ALT、AST活力均显著升高（P<0.01）；醋炙品组ALT、AST活力显著升高（P<0.01）；酒炙品组ALT、AST活力均显著升高（P<0.01）；炒焦品组ALT、AST活力均显著升高（P<0.01）；盐炙品组ALT、AST活力均显著升高（P<0.05）。与生品组相比，醋炙品组ALT值显著降低（P<0.01）；炒焦品组ALT值显著降低（P<0.05）；盐炙品组ALT显著降低（P<0.05），盐炙品组AST值显著降低（P<0.01）。
2.7.2　肝组织病理切片检查
光镜下观察如插页Ⅲ图7，正常组肝组织结构正常，无变性、坏死及炎症细胞浸润。生品组肝小叶结构保存，肝细胞弥漫中度水变性，可见个别点状坏死灶，汇管区少量炎症细胞浸润。醋炙品、炒焦品、盐炙品组肝小叶结构保存，肝细胞弥漫中度水变性，可见个别点状坏死灶，汇管区无炎症细胞浸润。酒炙品组肝小叶结构保存，肝细胞弥漫中度水变性，可见个别点状坏死灶，汇管区少量炎症细胞浸润。
2.7.3　SOD、GSH-Px、CAT、XOD活性检测及MDA、GSH含量测定
由表4结果可知，与正常组相比，川楝子生品与各炮制品的小鼠肝组织SOD活力均无明显差异。川楝子生品MTD组、酒炙品MTD组、酒炙品0.852MTD组、炒焦品0.85MTD组的小鼠肝组织GSH-Px活力显著降低（P<0.05或P<0.01），醋炙品和盐炙品GSH-Px活力无明显差异。川楝子生品MTD、0.852MTD组，醋炙品MTD、0.85MTD组，酒炙品MTD、0.852MTD组，炒焦品MTD组，盐炙品MTD组的小鼠肝组织CAT活力显著降低，其他组均无明显差异。川楝子生品MTD、0.852MTD、0.854MTD组，酒炙品MTD，炒焦品MTD组的小鼠肝组织XOD活力显著升高，醋炙品和盐炙品各组XOD活力均无明显差异。由表5结果显示，与正常组相比，川楝子生品MTD组、醋炙品MTD组、酒炙品MTD组的小鼠肝组织MDA含量显著升高（P<0.05或P<0.01），其他各组MDA均无明显差异。川楝子生品MTD组、炒焦品MTD组小鼠肝组织GSH含量显著降低，其他各组GSH含量均无明显差异。该结果提示川楝子生品致小鼠肝毒性的损伤机制与其抗氧化防御系统遭到破坏，清除自由基的能力降低，继而导致自由基在体内的过量堆积诱导细胞膜脂质过氧化反应有关。
3　讨论
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在临床用药历程中，中药肝毒性损伤（drug-inducedliverinjury，DIL）是一种临床上常见的药物不良反应，它的毒性在药物性靶器官毒性中排第2位，仅次于心脏毒性[19]。肝脏作为药物解毒的一个重要靶器官，承受着药物产生的毒性，随着中药在临床上的广泛应用，以及近年来中药毒理学研究的不断深入，使用中药导致的肝损伤问题报道越来越多，这一点引起了人们的高度重视，中药导致肝损伤问题已经成为了在中药使用过程中不得不注意的问题。药物的肝毒性研究在药物发现早期评价阶段起着至关重要的作用，而网络药理学结合分子对接方法已成为中药成分-靶点-机制挖掘研究的一项重要技术[20]。
目前，基于炮制减毒原理考察川楝子肝毒性的研究较少[21-22]，为深入探讨川楝子的物质基础及其机制靶点，本研究利用网络药理学方法预测其活性成分和作用靶点，并采用分子对接技术进行验证，结合炮制减毒原理为后续的研究提供基础。网络药理学作为中药成分靶点挖掘的新学科，可从系统和整体的角度衡量药物对生物分子网络的调节作用，具有系统性、相关性和可预测性等优势[23]。其与中药通过多成分、多靶点、多通路治疗疾病的理论一致。分子对接通过虚拟筛选技术寻找药物靶点，可高效、直观地展示药物与靶点间的作用关系[24]。因此，本研究运用网络药理学结合分子对接探讨川楝子毒性作用的物质基础及靶点通路具有一定的可行性。网络药理学得到川楝子致肝毒性的主要活性成分为苦楝子酮、苦楝素、印楝波为定A等。有研究表明，苦楝素能够通过触发PERK-eIF2α-ATF4介导的铁死亡诱导肝毒性[25]。PPI网络分析发现TP53、IL-6、CASP3、ESR1、HIF1A等是川楝子致肝毒性核心靶点。有研究显示，TP53此类关键信号分子的蛋白表达水平在肝毒性发生后显著升高[26]。有研究者发现克银丸潜在肝毒性机制可能是通过IL-17和细胞凋亡途径调节IL-6、肿瘤坏死因子α（TNF-α）、CASP3和CASP8靶点，从而引起炎症、因子过度释放和肝细胞坏死[27]。GO、KEGG富集分析显示，川楝子可通过干预多条炎症、糖脂代谢、细胞增殖凋亡信号通路导致肝毒性。包括TNF、PI3K-Akt、IL-17、HIF-1、细胞凋亡等信号通路。有研究通过斑蝥素处理大鼠肝脏中的免疫荧光染色发现了TNF-α、IkappaB激酶-α（IKK-α）和半胱天冬酶3的共表达[28]。
ALT、AST是两种氨基转移酶，升高主要反映肝细胞损伤，ALT、AST活力的增加与肝毒性损伤程度呈正相关，是肝毒性评价的一级指标。除此之外，氧化损伤机制也是中药致肝毒性损伤的主要机制。SOD、GSH-Px、CAT是机体的内源性抗氧化剂，其活力降低将造成机体抗氧化能力的下降使自由基清除障碍，从而导致自由基堆积造成肝细胞损伤。MDA是脂质过氧化反应链中的终产物，其高低间接反映了机体细胞受自由基攻击的严重程度。XOD是体内核酸代谢中重要酶，是鉴定肝细胞性黄疸及阻塞性黄疸的主要指标。体内实验研究发现，生品可引起明显的肝毒性，呈一定的剂量依赖性，醋炙品、炒焦品、盐炙品的毒性剂量范围缩小，肝组织病理损伤程度减轻，均可在不同程度下降低生品造成的过氧化脂质损伤，从而达到炮制减毒的效果，但研究发现川楝子经酒炙后肝毒性作用反而增强，不适合作为川楝子炮制减毒的处理方式。
综上所述，在氧化损伤的基础上，川楝子生品能导致肝损伤，醋炙、炒焦、盐炙后可降低川楝子肝损伤，可能通过减少炎症因子的释放，降低炎症反应减轻肝脏损伤，抑制肝细胞凋亡，达到炮制减毒目的，其中以醋炙品的减毒效果更为显著，但经酒炙的川楝子肝毒性明显高于生品，基于网络药理学和分子对接的预测结果进行研究，为后续明确川楝子炮制减毒原理奠定基础。
基金
收起
  • 国家中医药管理局“中药炮制传承技术传承基地”项目(桂中医药发〔2022〕9号)
  • 广西中医药大学桂派杏林拔尖人才资助项目(桂中医大学〔2022〕23号)
  • 广西中医药大学桂派中医药传承创新团队项目(2022B005)
  • 广西中医药大学硕士研究生科研创新重点项目(YCSZ2018015)
文献
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2025年第43卷第4期
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doi: 10.13193/j.issn.1673-7717.2025.04.001
  • 首发时间：2026-10-30
  • 出版时间：2025-04-10
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基金
国家中医药管理局“中药炮制传承技术传承基地”项目(桂中医药发〔2022〕9号)
广西中医药大学桂派杏林拔尖人才资助项目(桂中医大学〔2022〕23号)
广西中医药大学桂派中医药传承创新团队项目(2022B005)
广西中医药大学硕士研究生科研创新重点项目(YCSZ2018015)
作者信息
    广西中医药大学药学院，广西　南宁　530200

通讯作者:

杨柯（1975-），男（侗族），广西柳州人，教授，硕士研究生导师，硕士，研究方向：中药抗炎免疫药理。E-mail：。
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https://castjournals.cast.org.cn/joweb/zhzyyxk/CN/10.13193/j.issn.1673-7717.2025.04.001
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2种不同金属材料的力学参数
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Family		 属数
Number of
genus		 种数
Number of
species		 占总种数比例
Percentage of
total species (%)		 属
Genus		 种数
Number of
species		 占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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