目的:探讨铁凋亡相关基因在食管鳞状细胞癌(ESCC)铁凋亡调控中的作用。
方法:ESCC数据集GSE161533和GSE20347从基因表达Omnibus(GEO)下载,以使用R软件鉴定差异表达基因(DEGs)。通过将DEGs与来自FerrDb的铁凋亡相关基因相交获得的ESCC铁凋亡相关基因使用GO和KEGG分析进行分析,蛋白质-蛋白质相互作用(PPI)网络分析,并通过Cytoscape进行核心基因鉴定。使用TCGA数据库验证鉴定的铁凋亡抑制基因,用RT-qPCR检测其在培养的正常食管癌细胞和ESCC细胞中的表达水平。六个铁凋亡抑制基因(RRM2,GCLC,TFRC,TXN,SLC7A11和EZH2)在ESCC细胞中被siRNA下调,用CCK8法和流式细胞术评估细胞增殖和凋亡的变化;进行Western印迹以检查细胞铁凋亡进程的变化。
结果:我们共鉴定了58个ESCC铁凋亡相关基因,涉及谷胱甘肽跨膜转运等生物过程,铁离子传输,细胞凋亡和铁死亡,谷胱甘肽代谢,和抗叶酸的途径。PPI网络包括54个节点和74条边,聚类系数为0.522,PPI富集P<0.001。Cytoscape鉴定出6个核心铁凋亡抑制基因(RRM2,TFRC,TXN,EZH2、SLC7A11和GCLC),在TCGA数据集中的ESCC组织和ESCC细胞系中高度表达。在ESCCTE1细胞中下调这些基因显著抑制细胞增殖,促进细胞凋亡,降低了铁凋亡标志物GPX4和FIH1的表达水平,并增加了ACSL4的表达。
结论:在ESCC中铁凋亡抑制基因的高表达可能导致铁凋亡进程的停滞,从而促进肿瘤的发展。抑制这些基因可以恢复铁凋亡,促进细胞凋亡,提示其作为ESCC潜在治疗靶点的价值。
OBJECTIVE: To explore the role of ferroptosis-related genes in regulating ferroptosis of esophageal squamous cell carcinoma (ESCC).
METHODS: ESCC datasets GSE161533 and GSE20347 were downloaded from the Gene Expression Omnibus (GEO) to identify the differentially expressed genes (DEGs) using R software. ESCC ferroptosis-related genes obtained by intersecting the DEGs with ferroptosis-related genes from FerrDb were analyzed using GO and KEGG analyses, protein-protein interaction (PPI) network analysis, and core gene identification through Cytoscape. The identified ferroptosis suppressor genes were validated using TCGA database, and their expression levels were detected using RT-qPCR in cultured normal esophageal cells and ESCC cells. Six ferroptosis suppressor genes (RRM2, GCLC, TFRC, TXN, SLC7A11, and EZH2) were downregulated with siRNA in ESCC cells, and the changes in cell proliferation and apoptosis were assessed with CCK8 assay and flow cytometry; Western blotting was performed to examine the changes in ferroptosis progression of the cells.
RESULTS: We identified a total of 58 ESCC ferroptosis-related genes, which involved such biological processes as glutathione transmembrane transport, iron ion transport, and apoptosis and the ferroptosis, glutathione metabolism, and antifolate resistance pathways. The PPI network included 54 nodes and 74 edges with a clustering coefficient of 0.522 and PPI enrichment P<0.001. Cytoscape identified 6 core ferroptosis suppressor genes (RRM2, TFRC, TXN, EZH2, SLC7A11, and GCLC), which were highly expressed in ESCC tissues in the TCGA dataset and in ESCC cell lines. Downregulating these genes in ESCC TE1 cells significantly inhibited cell proliferation, promoted cell apoptosis, reduced the expression levels of ferroptosis markers GPX4 and FIH1, and increased the expression of ACSL4.
CONCLUSIONS: High expression of ferroptosis suppressor genes in ESCC may cause arrest of ferroptosis progression to facilitate tumor development, and inhibiting these genes can restore ferroptosis and promote cell apoptosis, suggesting their value as potential therapeutic targets for ESCC.