Senegenin mitigates neuroinflammation, pyroptosis, and apoptosis in cerebral ischemia via inhibiting STING and downstream inflammatory pathway.

Ischemic stroke continues to be a major cause of mortality and persistent disability, with neuroinflammation at the central stage of cell death signaling. The stimulator of interferon genes (STING) pathway is emerging as a central driver of microglial activation and inflammatory damage. However, therapeutic strategies targeting this pathway are limited. We investigated the neuroprotective effects of senegenin, a bioactive natural compound, in a rat middle cerebral artery occlusion/reperfusion (MCAO/R) model and N9 microglia subjected to oxygen-glucose deprivation/reoxygenation (OGD/R). Behavioral, histological, and biochemical analyses were performed to assess neurological outcomes, infarct volume, microglial activation, and neuroinflammatory response. Mechanistic studies evaluated the effects of senegenin on STING-TBK1-IRF3 signaling, NFκB-dependent NLRP3 inflammasome activation, pyroptosis, and apoptosis. Molecular docking, dynamics simulations, and pharmacological validation with the STING agonist DMXAA were used to confirm direct STING inhibition. Senegenin treatment significantly improved neurological outcomes, decreased infarct volume, and preserved cortical and hippocampal neurons. It attenuated oxidative stress, reduced DNA damage, and inhibited microglial activation. Mechanistically, senegenin suppressed STING activation and downstream phosphorylation of TBK1 and IRF3, blocked NF-κB/NLRP3-mediated pyroptosis, and inhibited apoptotic death by modulating Bcl2 and BAX expression. Molecular docking predicted stable binding of senegenin to STING, and DMXAA experiments confirmed direct inhibition of STING signaling as the mechanistic basis of its neuroprotective effects. This study demonstrates that senegenin confers potent neuroprotection in ischemic stroke by attenuating regulated cell death pathways through direct inhibition of STING, highlighting its ability as a promising therapeutic candidate for STING-targeted interventions in ischemic stroke and related neuroinflammatory disorders.