Identification of differentially expressed ferroptosis-related hub genes and their association with cigarette smoke-induced chronic obstructive pulmonary disease by bioinformatic analysis

Chronic Obstructive Pulmonary Disease (COPD) is a complex and progressive respiratory disorder, primarily resulting from exposure to cigarette smoke. The disease is characterized by chronic inflammation, oxidative stress, and irreversible destruction of lung tissue. Recent studies have revealed that ferroptosis, a regulated form of cell death driven by iron-dependent lipid peroxidation, plays a significant role in COPD pathogenesis. The aim of this study was to identify ferroptosis-related genes (FRGs) involved in cigarette smoke-induced COPD, and to investigate the expression of these genes in vitro. Initially, an integrative bioinformatic analysis was performed that was followed by in vitro cigarette smoke extract (CSE) exposure and expression analysis of target genes. The datasets used in this study were obtained from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were identified through dataset analysis, and ferroptosis-related genes were extracted using the FerrDb database gene list. The MCODE method, which was integrated into the Metascape online tool, was employed to identify hub genes. Subsequently, protein-protein interaction (PPI) analysis, functional enrichment analysis, and miRNA-gene interaction studies were conducted for the hub genes. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to analyse the expression of these hub genes after in vitro CSE exposure. Based on the analysis of multiple datasets and the FerrDb database, 20 FRGs involved in cigarette smoke-induced COPD were identified after removing redundant genes. MCODE analysis identified MDM2, EGFR, SRC, BRD4, GDF15, TP63, and CD44, as hub genes. Functional enrichment analysis revealed a strong association between these genes and ferroptosis-related biological pathways, including fatty acid and sulphur compound regulation that were mediated through the TP53 signaling pathway. miRNA-gene interaction analysis suggested that hsa-mir-203a-3p, hsa-mir-155-5p, and hsa-mir-34a-3p may regulate these genes. In vitro validation studies demonstrated that 24 hours incubation with CSE led to a significant reduction in the expression of six of the seven hub genes, except MDM2 gene in primary human bronchial epithelial cells (HBEC). However, CSE exposure did not cause any significant changes in the expression of genes studied in BEAS-2B cell line. In conclusion, we demonstrated the involvement of six ferroptosis-related hub genes in the pathogenesis of CSE-induced COPD by both bioinformatic tools and the validation by qRT-PCR. Our findings suggest that these genes may serve as potential targets for the diagnosis and treatment modalities of cigarette smoke-associated COPD; however, further studies are required.