目的 深度分析骨关节炎(osteoarthritis,OA)软骨细胞分化的单细胞转录组数据,探究其关键性靶基因及分子机制,为临床靶向治疗骨关节炎提供分子理论依据.方法 从GEO数据库下载数据集(GSE104782),包含10个骨关节炎样本及1 464个软骨单细胞测序结果,运用R语言分析软骨细胞分化中关键Marker基因,并采用质控和数据过滤、PCA、TSNE、细胞轨迹、GO、KEGG信号通路及蛋白互助网络分析揭示软骨细胞分化机制.结果 该数据集共包含基因17 380种、20个主成分、9种细胞类型、1 886个Marker基因及6种分化途径,其中软骨祖细胞为软骨最早分化细胞.GO分析涉及软骨发育、结缔组织发育等生物过程;涉及含胶原蛋白细胞外基质、内质网内腔等细胞组分;涉及肝素结合、糖胺聚糖结合等分子功能.KEGG通路分析涉及蛋白质消化吸收、ECM受体相互作用、人乳头瘤病毒感染、补体与凝血级联反应、PI3K-Akt信号通路等信号通路.运用Cytoscape 3.7.2软件获得5个关键Marker基因(COL1A1、COL2A1、VEGFA、COL1A2、DCN).结论 运用单细胞转录组测序发现骨关节炎软骨细胞共有6种分化途径及多种Marker基因的潜在特征,进一步从单细胞角度阐述骨关节炎的发病机制.
Objective To investigate the key target genes and molecular mechanisms of osteoarthritis chondrocyte differentiation,and provide molecular theoretical basis for clinical targeted therapy of osteoarthritis patients.Methods The GSE 104782 microarray dataset was downloaded from the GEO database,including the single-cell sequencing results of 1 464 chondrocytes from 10 osteoarthritis patients,and the R language tool was used to conduct in-depth analysis to obtain the key Marker genes in the process of chondrocyte differentiation.And data filtering,PCA analysis,TSNE analysis,cell trajectory analysis,GO enrichment analysis,KEGG signaling pathway analysis and protein interaction network analysis further confirmed the mechanism of chondrocyte differentiation.Results With the continuous deepening of sequencing,the gene features became more abundant;the data set contained 17 380 genes,20 principal components,9 cell types,1 886 Marker genes and 6 differentiation pathways,including the earliest differentiation of cartilage cell.GO enrichment analysis found that biological process(BP)involved cartilage development,connective tissue development;cellular component(CC)involved collagen-containing extracellular matrix,endoplasmic reticulum lumen;molecular function(MF)involved heparin binding,glycosaminoglycan binding.KEGG pathway analysis:Mainly involved in protein digestion and absorption,ECM receptor interaction,human papillomavirus infection,focal adhesion,proteoglycan in cancer,complement and coagulation cascade,PI3K-Akt signaling pathways,mineral absorption and other signal pathways.Cytoscape 3.7.2 software was used to obtain 5 key target genes(COL1A1,COL2A1,VEGFA,COL1A2,DCN)in marker.Conclusion Using single-cell transcriptome sequencing,we found that osteoarthritic chondrocytes have six differentiation pathways and potential features of multiple marker genes,which further elucidated the pathogenesis of osteoarthritis from a single-cell perspective.