Xylazine‐Adulterated Fentanyl‐Induced Brain Toxicity: Mechanism Elucidation via Network Toxicology and Molecular Docking Approaches

ABSTRACT Xylazine‐adulterated fentanyl (FAAX) is emerging as a public health concern, causing life‐threatening brain injuries that account for overdose deaths; however, the underlying mechanisms are far from illustrated. The present study aimed to explore and analyze the molecular mechanisms of FAAX‐induced brain toxicity through network toxicology approaches and in vitro validation. 294 potential targets were identified, and protein–protein interaction (PPI) analysis ranked six proteins as core targets, among which JAK2 and PI3K were emphasized. Molecular docking revealed that both fentanyl and xylazine bind JAK2 and PI3K with high affinity. In SH‐SY5Y cells treated with FAAX (fentanyl 40 μM, xylazine 160 μM, 24 h), RT‐qPCR and Western blotting confirmed marked upregulation of JAK2 and PI3K. Collectively, we determined JAK2 and PI3K as the key targets in FAAX‐induced brain toxicity. Our findings not only offer a novel perspective for future research but also provide a foundation for future translational studies aimed at mitigating the neurological consequences of FAAX exposure.