Dehydroabietic acid protects against cerebral ischaemia–reperfusion injury by modulating microglia‐mediated neuroinflammation via targeting PKCδ

Abstract Background and Purpose Cerebral ischaemia–reperfusion injury (CIRI) is a major contributor to global morbidity and mortality, although its underlying mechanisms remain only partly understood. Emerging evidence indicates that inhibiting microglia‐mediated neuroinflammation would be an effective therapeutic approach for CIRI, and pharmacological interventions targeting this pathway hold significant therapeutic promise. This study aimed to identify a potent anti‐inflammatory drug from a natural compound library as a potential treatment for CIRI. Experimental Approach We used oxygen–glucose deprivation/reperfusion (OGD/R) and middle cerebral artery occlusion in male C57BL/6 mice to evaluate the efficacy of DHA in neurological deficits and the anti‐inflammatory effects. Using BV2 cells and murine brain tissue, liquid chromatography–tandem mass spectrometry was used to identify potential molecular targets of DHA, followed by bio‐layer interferometry, molecular docking, molecular dynamics simulations and cellular thermal shift assays to validate DHA's binding interactions with protein kinase C delta (PKCδ). Key Results DHA decreased production of pro‐inflammatory cytokines following OGD/R, thereby inhibiting microglia‐mediated neuroinflammation to protect neurons and reducing brain infarct size and improving neurological outcomes. Mechanistically, DHA directly bound to PKCδ, inhibiting its phosphorylation and downstream NF‐κB signalling. This binding interaction involved TRP 55 and LEU 106 on PKCδ, as confirmed by molecular docking and other biophysical techniques. Conclusion and Implications DHA specifically interacts with PKCδ, preventing its phosphorylation induced by ischaemia–reperfusion injury. These results suggest that DHA is a novel inhibitor of PKCδ and provide solid experimental foundations for using DHA in treating neuroinflammation‐related conditions, such as CIRI.