Integrated bioinformatic analysis and experimental validation for exploring the key immune checkpoint of COPD

There is growing evidence indicating immune inflammation is a key factor in the progression of chronic obstructive pulmonary disease (COPD). Immune checkpoints (ICs) are crucial targets for modulating the functional activation and differentiation of immune cells, particularly in relation to immune inflammation and the regulation of T cell activation and exhaustion. However, the precise mechanisms of ICs in COPD remain understood. COPD datasets were obtained from the Gene Expression Omnibus (GEO) and analyzed using GEO2R and Limma to identify differentially expressed genes. LASSO regression was then applied to screen ICs closely associated with COPD. Finally, target genes were selected based on gene expression profiles. Gene ontology (GO), immune infiltration analysis, and gene set enrichment analysis (GSEA) were utilized to assess the relationship between IC genes (ICGs) and immune cells. Subsequently, tobacco-exposed mice, anti-Tim3-treated mice, and HAVCR2-knockout mice were generated, with flow cytometry being used to confirm the results. Through the analysis of GSE38974 and LASSO regression, five ICGs were identified. Subsequent validation using GSE20257 and GSE76925 confirmed these findings. Gene expression profiling highlighted HAVCR2 as having the strongest correlation with COPD. Further investigation through immune infiltration analysis, GO, and GSEA indicated a link between HAVCR2 and CD8+ T cells in COPD. Flow cytometry experiments demonstrated high Tim3 expression in CD8+ T cells of mice exposed to tobacco, promoting Tc1 and inhibiting Tc17, thus affecting CD8+ Tem activation and CD8+ Tcm formation, leading to an immune imbalance within CD8+ T cells. Prolonged exposure to tobacco upregulates Tim3 in CD8+ T cells, triggering its regulatory effects on Tc1/Tc17. Knocking out HAVCR2 further upregulated the expression of CD8+ Tem while suppressing the expression of CD8+ Tcm, indicating that Tim3 plays a role in the activation and differentiation of CD8+ T cells in the context of tobacco exposure.