Identification of key genes related to metabolic cell death in hepatic ischemia-reperfusion injury from transcriptome data and mechanism research using single-cell data.

Background: Ferroptosis and cuproptosis are closely associated with hepatic ischemia-reperfusion injury (HIRI). However, the significance of metabolic cell death-related genes (MRGs) in HIRI still awaits exploration. This study examined the molecular mechanisms through which key genes contribute to metabolic cell death in HIRI. Methods: GSE12720, GSE14951, and GSE171539 datasets and 478 MRGs were included. First, candidate genes were screened through differential expression analysis combined with MRGs. Then, key genes were identified by using machine learning algorithms combined with expression verification. Subsequently, the analyses included constructing and evaluating nomograms, conducting functional enrichment studies, characterizing immune infiltration, building regulatory networks, performing drug prediction, and executing molecular docking. Importantly, single-cell analysis was conducted to identify key cell populations Finally, expression levels of key genes in animal samples were determined by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Results: The analysis identified ATF3, TNFAIP3, IL1B, and KDM6B as central genes. The nomogram indicated that these four key genes could well predict the occurrence of HIRI. Functional enrichment analysis revealed significant associations of ATF3, TNFAIP3, and KDM6B with olfactory transduction pathways. The key genes were positively linked to most differential immune cells, and ATF3 had the most significant positive relation to activated CD4 T cells. The binding energies of molecular docking between key genes and corresponding drugs were all less than -5 kcal/mol. Mononuclear phagocytes were identified as key cells, and the expressions of ATF3, IL1B, and KDM6B had dynamic and non-linear change characteristics during the differentiation of mononuclear phagocytes. RT-qPCR results demonstrated that ATF3, TNFAIP3, IL1B, and KDM6B were up-regulated in HIRI samples, consistent with the results in the GSE12720 and GSE14951 datasets. Conclusion: In HIRI pathogenesis research, ATF3, TNFAIP3, IL1B and KDM6B were validated as core regulators of metabolic cell death, offering critical targets for mechanistic investigation.