基于生物信息学分析筛选阳虚质类风湿关节炎的差异表达基因及潜在干预中药

Bioinformatics Analysis of Differentially Expressed Genes and Potential Chinese Medicine Screening for Rheumatoid Arthritis with Yang Deficiency Constitution

目的:利用生物信息学分析筛选阳虚质类风湿关节炎(rheumatoid arthritis,RA)的差异表达基因(differentially expressed genes,DEGs),并对其干预中药进行预测,为深入研究阳虚质RA相关病理机制及干预中药的用药规律提供依据。方法:从基因表达综合数据库(gene expression omnibus,GEO)获取中医阳虚质数据集GSE87474和RA相关数据集GSE90081、GSE193193。合并GSE90081和GSE193193数据集形成RA合并数据集,并用替代变量分析(surrogate variable analysis,SVA)包消除批次效应。利用微阵列数据线性模型(linear models for microarray data,limma)包分别筛选GSE87474和RA合并数据集的DEGs,取交集获取阳虚质RA相关基因。然后,利用机器学习算法,从阳虚质RA相关基因中鉴定关键基因,绘制其受试者工作特征曲线,计算曲线下面积(area under curve,AUC)值,并对每个关键基因进行基因集富集分析(gene set enrichment analysis,GSEA)。同时,通过医学信息检索平台(Coremine Medical)数据库预测干预阳虚质RA的中药,并收集其性味归经等信息。结果:筛选出264个RA相关DEGs,455个阳虚质DEGs,取交集后获得17个(上调15个,下调2个)阳虚质RA相关基因。依据随机森林(random forest,RF)模型算法共鉴定出5个关键基因,即肾上腺髓质素(adrenomedullin,ADM)、分泌性白细胞肽酶抑制剂(secretory leukocyte peptidase inhibitor,SLPI)、膜联蛋白A3(annexin A3,ANXA3)、G蛋白信号调节器18(regulator of G protein signaling 18,RGS18)、C-C基序趋化因子配体4(C-C motif chemokine ligand 4,CCL4)。这5个关键基因的AUC均>0.750。GSEA结果显示,5个关键基因功能富集共同集中在16条通路,涉及转化生长因子-β(transforming growth factor-β,TGF-β)信号通路、氧化磷酸化、谷胱甘肽代谢等。中药预测结果显示:潜在干预阳虚质RA的中药多归肝、肺、脾胃、肾经,药性以温为主,药味以苦、甘、辛多见。结论:ADM、SLPI、ANXA3、RGS18、CCL4可能为阳虚质RA的潜在关键基因,可调控TGF-β信号通路、氧化磷酸化、谷胱甘肽代谢等途径介导阳虚质RA的病理过程;干预阳虚质RA的中药大多归肝、肺、脾胃、肾经,性味多为温、苦、甘、辛。

Objective:To screen differentially expressed genes(DEGs)in rheumatoid Arthritis(RA)with Yang deficiency constitution by bioinformatics analysis,and to predict potential Chinese medicine for the intervention of it so as to provide a basis for further study of related pathological mechanism and medication rules.Methods:The TCM Yang deficiency constitution dataset GSE87474 and the RA related datasets GSE90081 as well as GSE193193 were obtained from the gene expression omnibus(GEO)database.The GSE90081 and GSE193193 datasets were combined to form the RA pooled dataset,and the surrogate variable analysis(SVA)package was used to eliminate batch effects.DEGs in the combined GSE87474 and RA pooled datasets were respectively screened using linear models for microarray data(limma)package,and the intersection was taken to obtain the genes related to RA with Yang deficiency constitution,from which the key genes were identified by machine learning algorithm and then used to draw the receiver operating characteristic curve as well as calculating the area under curve(AUC)value.Gene set enrichment analysis(GSEA)was performed for each key gene.In addition,Coremine Medical database was used to predict the Chinese medicine for RA with Yang deficiency constitution,and the information of the properties,tastes and Meridian tropism was collected.Results:A total of 264 DEGs related to RA and 455 DEGs related to Yang deficiency constitution were screened,and 17 DEGs related to RA with Yang deficiency constitution(15 up-regulated and 2 down-regulated)were obtained after taking the overlap.A total of 5 key genes were identified based on the random forest(RF)model algorithm,including adrenomedullin(ADM),secretory leukocyte peptidase inhibitor(SLPI),annexin A3(ANXA3),regulator G protein signaling 18(RGS18)and C-C motif chemokine ligand 4(CCL4).The AUCs of these five key genes were all greater than 0.750.The results of GSEA showed that the functional enrichment of 5 key genes was concentrated in 16 pathways,involving transforming growth factor-β(TGF-β)signaling pathway,oxidative phosphorylation,glutathione metabolism,etc.The prediction results of Chinese medicine showed that the potential Chinese medicinal for the intervention of RA with Yang deficiency constitution mostly entered the Liver,Lung,Spleen and Stomach as well as Kidney meridians.The medicinal property were mainly warm,and the tastes were mostly bitter,sweet,acrid or pungent-spicy.Conclusion:ADM,SLPI,ANXA3,RGS18 and CCL4 may be the potential key genes of RA with Yang deficiency constitution,which can mediate the pathological process of RA with Yang deficiency constitution by regulating TGF-βsignaling pathway,oxidative phosphorylation and glutathione metabolism.Most of the Chinese medicines that treat RA with Yang deficiency constitution enter the Liver,Lung,Spleen and Stomach as well as Kidney meridians,with warm property,bitter,sweet,acrid or pungent-spicy tastes.