Extracellular cold-inducible RNA-binding protein drives NLRP3-mediated pyroptosis in acute ischemic stroke.

Acute ischemic stroke (AIS) triggers rapid sterile inflammation that worsens secondary brain injury. Damage-associated molecular patterns (DAMPs) from stressed/necrotic cells activate inflammasomes and pyroptosis. Cold-inducible RNA-binding protein (CIRP), a stress-responsive DAMP linked to maladaptive inflammation, has unclear clinical relevance to ischemic stroke and mechanisms in post-ischemic neuroinflammation. We measured serum CIRP, neuronal injury markers (neuron-specific enolase [NSE] and S100 calcium-binding protein β [S100β]), and pyroptosis mediators (NOD-like receptor family pyrin domain-containing 3 [NLRP3] inflammasome components, gasdermin D [GSDMD], and interleukin-1β [IL-1β]) in patients with AIS and in controls and analyzed correlations. In rat middle cerebral artery occlusion (MCAO) models, we profiled CIRP, neuroinflammation, and pyroptosis in the penumbra and perturbed CIRP by knockout or the inhibitory peptide C23. Outcomes were infarct volume, neurological scores, cytokines, and activation of the NLRP3-caspase-1-GSDMD axis. Serum CIRP was elevated in AIS and correlated with injury and pyroptosis markers. CIRP was upregulated in ischemic brain with inflammasome activation and cytokine release. CIRP knockout or C23 reduced infarct size, improved function, and suppressed pyroptosis and neuroinflammation. Notably, our clinical cohort comprised patients who did not receive reperfusion therapies, highlighting CIRP as a biomarker-linked inflammatory driver and a tractable therapeutic node for late-presenting AIS patients ineligible for intravenous thrombolysis. Overall, CIRP amplifies ischemic injury via NLRP3 inflammasome-mediated pyroptosis, while C23 confers in vivo neuroprotection, nominating CIRP as a tractable target in AIS particularly late-presenting, non-thrombolysed patients.