淫羊藿通过ICA-miR-335-5p轴缓解骨质疏松症的生物信息学分析及实验研究  

Bioinformatics Analysis and Experimental Study of Yinyanghuo(Epimedium)in Alleviating Osteoporosis Through ICA-miR-335-5p Axis

作　　者：金垚 徐鹏辉 卞泗善[2] 滕加文[2] JINYao;XU Penghui;BIAN Sishan;TENG Jiawen(The First Clinical Medical College of Shandong University of Chinese Medicine,Jinan Shandong 250014,China;Affiliated Hospital of Shandong University of Chinese Medicine,Jinan Shandong 250014,China)

机构地区：[1]山东中医药大学第一临床医学院,山东济南250014 [2]山东中医药大学附属医院,山东济南250014

出　　处：《中医药导报》2023年第2期1-7,13,共8页Guiding Journal of Traditional Chinese Medicine and Pharmacy

基　　金：山东省自然科学基金面上项目(ZR2020MH359)。

摘　　要：目的:通过生物信息学结合动物实验探讨淫羊藿苷(ICA)上调miR-335-5p促进大鼠成骨分化缓解OP的分子机制。方法:利用TCMSP数据库筛选淫羊藿的主要活性成分及靶点,通过GEO数据库进行miR-335-5p的差异性分析,采用相应数据库预测miR-335-5p和OP的基因靶点,对淫羊藿、miR-335-5p和OP靶点求交集,筛选出淫羊藿干预miR-335-5p治疗OP的基因;利用Metascape数据库进行富集分析,并用String数据库构建PPI网络筛选出Hub基因后,在AutoDock Tools 1.5.6软件中进行分子对接,模拟药效结合过程;采用RT-qPCR测定miR-335-5p在OP患者骨组织、骨质疏松症大鼠骨组织的表达,并测定成骨诱导OP患者BMSCs分化14 d后的miR-335-5p表达,口服ICA和/或过表达miR-335-5p治疗骨质疏松症大鼠10周后,检测成骨基因的表达,HE染色观察大鼠股骨干骺端骨组织微观结构。结果:淫羊藿主要活性成分有23个、作用靶点有221个,淫羊藿调控miR-335-5p治疗OP的作用靶点有40个,富集分析涉及多条通路,分子对接结果显示淫羊藿苷与Hub蛋白均可形成稳定化合物发挥作用。实验研究发现,miR-335-5p在OP患者和OP大鼠的骨组织中低表达,ICA处理和/或过表达miR-335-5p可促进大鼠BMSCs成骨分化;HE染色显示造模大鼠经ICA处理和/或过表达miR-335-5p后骨小梁增多、排列紧密,部分新生骨组织形成,有效缓解了大鼠骨质疏松症。结论:淫羊藿苷可通过ICA-miR-335-5p轴促进大鼠成骨分化来缓解OP,为中药单体治疗OP提供了新的研究思路。Objective:To explore the molecular mechanism of icariin(ICA)up-regulating miR-335-5p to promote osteogenic differentiation in rats and alleviate OP by bioinformatics combined with animal experiments.Methods:The TCMSP database was used to screen the main active components and targets of Yinyanghuo(Epimedium),the GEO database was used to analyze the differences of miR-335-5p,and the corresponding database was used to predict the gene targets of miR-335-5p and OP,and the intersection of Yinyanghuo(Epimedium),miR-335-5p and OP targets was used to screen the genes of Yinyanghuo(Epimedium)interfering with miR-335-5p to treat OP.The Metascape database was used for enrichment analysis and the String database was used to construct a PPI network to screen out the Hub genes,then carry out molecular docking in AutoDock Tools 1.5.6 software to simulate the process of drug combination.RT-qPCR was used to determine the expression of miR-335-5p in the bone tissue of OP patients and osteoporotic rats bone tissue,and the expression of miR-335-5p after 14 days of differentiation of BMSCs induced by osteogenesis in OP patients.After 10 weeks of treatment with oral ICA and/or overexpression of miR-335-5p in osteoporotic rats,the expression of osteogenic genes was detected,and HE staining was used to observe the microstructure of bone tissue in the epiphysis of rat femoral shaft.Results:Yinyanghuo(Epimedium)had 23 main active components and 221 targets.Yinyanghuo(Epimedium)regulated 40 targets of miR-335-5p in the treatment of OP.The enrichment analysis involved multiple pathways.The molecular docking results showed that icariin and Hub protein can form stable compounds to play a role.The experimental study found that miR-335-5p was low expressed in the bone tissue of OP patients and OP rats,and ICA treatment and/or overexpression of miR-335-5p could promote the osteogenic differentiation of rat BMSCs.HE staining showed that after ICA treatment and/or overexpression of miR-335-5p,the bone trabeculae of the model rats increased and arr

关 键 词：骨质疏松症 淫羊藿 淫羊藿苷 生物信息学 BMSCS 成骨分化 大鼠 

分 类 号：R285.5[医药卫生—中药学]