不明原因复发性流产中铁死亡的生物信息学分析及验证

Bioinformatic analysis and validation of ferroptosis in unexplained recurrent spontaneous abortion

目的:探究铁死亡关键基因在不明原因复发性流产(URSA)发生发展中的作用,初步确定潜在生物标志物。方法:从基因表达综合(GEO)数据库下载数据集GSE26787,利用GEO2R筛选差异表达基因(DEGs);从FerrDb V2数据库下载铁死亡相关基因;DEGs与铁死亡基因取交集获得URSA铁死亡相关基因;利用DAVID数据库对URSA铁死亡相关基因进行基因本体(GO)富集及京都基因与基因组百科全书(KEGG)通路分析;利用String数据库和Cytoscape软件分析蛋白互作网络,筛选Hub基因;采用实时荧光定量PCR(RT-qPCR)检测Hub基因在人工流产组及URSA组患者蜕膜组织中mRNA的表达水平。结果:共筛选出55个URSA铁死亡基因;GO功能富集提示URSA铁死亡相关基因主要在自噬调节、RNA聚合酶Ⅱ启动子转录正调控、膜的整体组分、酶及p53蛋白受体结合富集。KEGG通路分析显示URSA铁死亡基因富集最明显的为FoXO信号通路。采用String数据库及Cytoscape软件筛选出的URSA铁死亡关键基因分别为EGFR、SRC、KRAS、MDM2。RT-qPCR检测结果显示,EGFR、SRC、MDM2在URSA组中的表达均低于人工流产组(均P<0.05);KRAS在URSA组和人工流产组中表达无统计学差异(P>0.05)。结论:铁死亡相关基因EGFR、SRC、MDM2可作为诊治URSA的潜在生物标志物。

Objective:To explore the the role of key genes of ferroptosis in the occurrence and development of unexplained recurrent spontaneous abortion(URSA),and to preliminarily identify the potential biomarkers.Methods:The GSE26787 data set was downloaded from Gene Expression Synthesis(GEO)database,and the differentially expressed genes(DEGs)were screened by GEO2R,obtaining ferroptosis-related genes from FerrDb V2 database.The ferroptosisrelated genes in URSA were obtained by intersection of DEGs and ferroptosis genes.The gene ontology(GO)enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis of ferroptosis-related genes in URSA were performed using DAVID database.The protein interaction network was analyzed using String database and Cytascape software to screen the Hub genes.Real-time fluorescence quantitative polymerase chain reaction(RT-qPCR)was used to detect the mRNA expression levels of Hub genes in the decidua tissues of patients in the induced abortion group and the URSA group.Results:A total of 55 ferroptosis-related genes in URSA were screened.GO functional enrichment analysis found that ferroptosis-related genes in URSA were mainly enriched in the regulation of macroautophagy,positive regulation of transcription from RNA polymeraseⅡpromoter,integral components of the membrane and enzyme and p53 receptor binding.The enrichment analysis of KEGG pathway showed that the most obvious enrichment of ferroptosis-related genes in URSA was the FoxO signaling pathway.String database and Cytoscape software were used to screen the key genes of ferroptosis EGFR、SRC、KRAS、MDM2 in URSA.The results of RT-qPCR showed that the expressions of EGFR,SRC and MDM2 in the URSA group were lower than those in the induced abortion group(all P<0.05).There was no statistically significant difference in the expression of KRAS between URSA group and induced abortion group(P>0.05).Conclusion:Ferroptosis-related genes EGFR,SRC and MDM2 can be used as potential biomarkers for diagnosis and treatment of URSA.