增强人尿源性干细胞干性的中药小分子活性成分筛选  

Screening of small molecular active components of traditional Chinese medicine to enhance the stemness of human urine-derived stem cells

作　　者：沈亮亮 李勇 周明 谯英固 孙震晓 SHEN Liang-liang;LI Yong;ZHOU Ming;QIAO Ying-gu;SUN Zhen-xiao(School of Life Sciences,Beijing University of Chinese Medicine,Beijing 102488,China)

机构地区：[1]北京中医药大学生命科学学院,北京102488

出　　处：《中国新药杂志》2020年第24期2839-2845,共7页Chinese Journal of New Drugs

基　　金：北京中医药大学2019年研究生自主课题资助项目(2019-JYB-XS-112)。

摘　　要：目的:以人尿源性干细胞(human urine-derived stem cells,hUSCs)Wnt信号通路中的糖原合成酶激酶(glycogen synthase kinase-3β,GSK-3β)为靶标,从中药系统药理学(traditional Chinese medicine systems pharmacology,TCMSP)数据库收录的中药小分子活性成分中筛选GSK-3β抑制剂,研究其是否具有增强hUSCs干性基因表达的作用。方法:参考京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)中人多能干细胞干性调控信号网络,以Wnt信号通路中的GSK-3β为靶点,利用分子对接技术对TCMSP数据库进行虚拟筛选,寻找能够增强hUSCs干性的中药小分子抑制剂;结合文献调研优化分子对接结果,通过MTT实验考察其对hUSCs细胞活力的影响,qRT-PCR实验考察其是否增强hUSCs干性基因的表达。结果:根据GSK-3β晶体结构复合的原配体对接结果,确定中药小分子活性成分的筛选规则为对接打分值高于阈值81.112(原配体打分值101.39的80%)且与受体GSK-3β的作用模式与原配体类似(与关键氨基酸残基ASP133和VAL135发生氢键作用);候选配体对接结果表明,黄芩素、白杨素、褪黑素与GSK-3β的对接打分值均高于阈值且与GSK-3β的作用模式均与原配体类似,可作为候选中药小分子活性成分;细胞实验结果表明,3种候选中药小分子活性成分在一定浓度范围内具有维持hUSCs细胞活力的作用,且均能显著上调hUSCs干性基因OCT4的表达水平,对MYC基因的表达水平无明显作用。结论:通过分子对接技术筛选到的针对GSK-3β的中药小分子抑制剂黄芩素、白杨素和褪黑素可以作为增强hUSCs干性的候选化合物,中药小分子活性成分有潜力被应用于hUSCs等人成体干细胞研究领域的药物研发。Objective:To screen GSK-3βinhibitors from the small molecular active components of traditional Chinese medicine included in traditional Chinese medicine systems pharmacology(TCMSP)database,using GSK-3β,a glycogen synthesis kinase in Wnt signaling pathway,as the target,and explore whether these inhibitors enhance the expression of stemness genes in human urine-derived stem cells(hUSCs).Methods:Referring to the signal network of stemness regulation of human pluripotent stem cells in Kyoto encyclopedia of genes and genomes(KEGG),molecular docking technology was used to search for GSK-3βsmall molecular inhibitors of traditional Chinese medicine in TCMSP database.The result of molecular docking was optimized according to previous literature reports,and the effect of the small molecular active components of traditional Chinese medicine on the viability of hUSCs and on the expression of the stemness genes of hUSCs were investigated by using MTT and qRT-PCR assays.Results:According to the docking result of the primary ligand complexed with the crystal structure of GSK-3β,it was determined that the screening rule of small molecule active components of traditional Chinese medicine was that its docking score was higher than the threshold value of 81.112(80%of the docking score of primary ligand of 101.39),and the mode of interaction with GSK-3βwas similar to that of the primary ligand(hydrogen bonding with the key amino acid residues ASP133 and VAL135).The docking results of candidate ligands showed that the docking scores of baicalein,chrysin and melatonin with GSK-3βwere all higher than the threshold value,and the modes of interaction with GSK-3βwere all similar to that of the primary ligand,indicating that they can become the candidate small molecule active components of traditional Chinese medicine.The results of cell experiment showed that the three candidates of small molecule active components of traditional Chinese medicine had the effect of maintaining the viability of hUSCs,up-regulating the expression leve

关 键 词：分子对接 干性 GSK-3β抑制剂 人尿源性干细胞 中药小分子活性成分 

分 类 号：R284[医药卫生—中药学] R932[医药卫生—中医学]