基于斑马鱼在体模型高效筛选补骨脂配伍减毒研究  

Analysis on the Compatibility of Psoralen Based on the Toxicity Evaluation of Zebrafish

作　　者：宁青 刘中秋[1] 韦英杰[2] 胡明 NING Qing;LIU Zhong-qiu;WEI Ying-jie;HU Ming(International Institute of Chinese Medicine Translation,Guangzhou University of Chinese Medicine,Guangzhou,510006,China;Jiangsu Academy of Chinese Medicine,Key Laboratory of Chinese Medicine Release System,State Administration of Traditional Chinese Medicine,Nanjing,210028,China)

机构地区：[1]广州中医药大学国际中医药医学转化研究所,广东广州510006 [2]江苏省中医药研究院,国家中医药管理局中药释药系统重点研究室,江苏南京210028

出　　处：《南京中医药大学学报》2021年第1期54-61,共8页Journal of Nanjing University of Traditional Chinese Medicine

基　　金：“重大新药创制”科技重大专项资助(2017ZX09301-056);国家自然科学基金(81573833)。

摘　　要：目的基于斑马鱼在体模型高效筛选补骨脂配伍减毒药味,探讨其减毒机制,为临床用药安全提供理论依据。方法采用受精后1~6 dpf斑马鱼,研究不同药味对斑马鱼的影响,观察鱼脏器行态/形态,计数死亡数,快速筛选确定安全浓度;以补骨脂素为代表毒性成分,观察不同药味与其配伍后对斑马鱼毒性的变化,筛选有效减毒的药味及成分;采用RNA-seq技术(RNA sequencing)对杜仲代表活性成分桃叶珊瑚苷配伍补骨脂素进行转录组测序,建立基因表达谱(DGE)文库,进行肝毒相关基因本体论(GO)功能注释和京都基因与基因组百科全书(KEGG)通路分析,筛选出肝脏相关通路中有代表性的差异表达基因,初步探讨桃叶珊瑚苷配伍减毒机制。结果通过筛选表明桃叶珊瑚苷可显著降低补骨脂对斑马鱼的肝脏毒性影响。RNA-Seq富集显示,加入桃叶珊瑚苷配伍后与原有的单一补骨脂素组相比,毒性、疾病(癌症)、免疫激活、化学致癌特征信号方面有明显的减少,神经营养因子、长寿调节、血小板激活信号通路增强。同时筛选出差异表达基因gpx1b、pgd、rrm2、gstp2、anpepb、cyp1a、prf1.5、nfκb2。结论利用斑马鱼在体快速评价的优势,实现基于体内过程的毒性评价,先从毒性明确的代表成分补骨脂素配伍可快速锁定减毒药味及成分,利用RNA-seq技术可进一步分析其配伍减毒的机制,该研究对其临床药味配伍减毒理论具有一定的意义。OBJECTIVE Based on the zebrafish in vivo model,the compatibility of psoralen for ameliorating medicinal property was singled out to explore its detoxification mechanism and provide a theoretical basis for Clinical medication safety.METHODS Using 1 to 6 dpf zebrafish after fertilization,the effects of different medicinal property on zebrafish was studied.the morphology/behavior of fish organs was observed and the number of deaths was counted.It determines the safe concentration by rapid screening.Psoralen is used as a representative toxic component to observe different medicinal property and their compatibility toxicity changes for zebrafish,that screen effective attenuated medicinal property and ingredients.RNA-seq was used to get the transcriptome of treating with aucubin(the representative and active ingredient of Eucommia)and psoralen to establish Gene expression profiling(DGE)library,Hepatotoxicity-related gene was selected by Gene Ontology(GO)functional annotation and Kyoto Encyclopedia of Genes and Genome(KEGG)pathway analysis.The differentially expressed genes in liver-related pathways were screened out and the detoxification mechanism of aucubin was preliminarily explored.RESULTS It shows that aucubin can significantly reduce the liver toxicity of psoralen on zebrafish with drug compatibility.Enrichment analysis of RNA-Seq showed that compared with the psoralen group,expression of gene in the toxicity,cancer,immune activation,and chemical carcinogenic signals were significantly reduced,neurotrophic factors,Longevity regulation,platelet activation signal pathway enhanced in addition of aucubin,among which the gene names as gpx1b,pgd,rrm2,gstp2,anpepb,cyp1a,prf1.5,nfκb2 about attenuating toxicity after compatibility,which is worthy of attention.CONCLUSION Taking advantage of the rapid evaluation of zebrafish in vivo,toxicity evaluation based on processes in vivo can be achieved.On the one hand,the compatibility of the representative component of psoralen with clear toxicity quickly targets the attenuated m

关 键 词：斑马鱼 补骨脂 配伍减毒 RNA-SEQ 

分 类 号：R285.5[医药卫生—中药学]