Integrated Multi-Omics Analysis Reveals IRF1-Driven Microglial PANoptosis via ZBP1 in Spinal Cord Injury.

BACKGROUND: Spinal cord injury (SCI) is a devastating condition with high disability rates. In the secondary injury phase, activated microglia play a central role by mediating the inflammatory response. Notably, pro-inflammatory forms of programmed cell death, including PANoptosis, can exacerbate inflammatory cascade. Although microglia-driven inflammation is well-characterized in SCI, it remains unclear whether microglia undergo PANoptosis. Furthermore, potential therapeutic agents targeting this specific process remain to be discovered in SCI. METHODS: We performed an integrated analysis of bulk RNA-seq and scRNA-seq to investigate the heterogeneity of programmed cell death and PANoptosis across different cell types. Candidate genes associated with high PANoptosis activity scores were identified using correlation analysis. Furthermore, four machine learning algorithms were applied to screen for core genes. By combining transcription factor (TF) activity prediction and Chip-atlas database analysis, we identified TFs that potentially regulate PANoptosis. Validation experiments were conducted at each analytical stage using animal and cellular models. RESULTS: Pyroptosis, apoptosis, and necroptosis, were persistently upregulated following SCI. Notably, the PANoptosis pathway was significantly enriched and predominantly localized to microglia. Further investigation identified ZBP1 as prominently upregulated PANoptosome sensor associated with microglial PANoptosis. Moreover, IRF1 was found to be enriched at the promoter region of Zbp1 and transcriptionally regulated its expression. Pharmacological inhibition of IRF1 was accompanied by reduced ZBP1 expression, suppression of PANoptosis-related execution markers, decreased TNF-α and IL-6 release, and attenuation of M1-like microglial polarization. CONCLUSION: These findings highlight the IRF1-ZBP1 axis as a regulatory mechanism of PANoptosis in microglia following SCI and suggest it as a potential therapeutic target to modulate neuroinflammation.