PKM2 promotes neutrophil activation and cerebral thrombo-inflammation: Therapeutic implications for ischemic stroke

Abstract

There is a critical need for cerebroprotective interventions to improve the suboptimal outcomes of patients with ischemic stroke treated with reperfusion strategies. We found that nuclear pyruvate kinase muscle 2 (PKM2), a modulator of systemic inflammation, was upregulated in neutrophils after the onset of ischemic stroke both in humans and in mice. Therefore, we determined the role of PKM2 in stroke pathogenesis utilizing murine models with preexisting comorbidities. We generated novel myeloid cell-specific PKM2-/- mice on wild-type (PKM2fl/flLysMCre+) and hyperlipidemic background (PKM2fl/flLysMCre+Apoe-/-). Controls were littermate PKM2fl/flLysMCre- or PKM2fl/flLysMCre-Apoe-/- mice. Genetic deletion of PKM2 in myeloid cells limited inflammatory response in peripheral neutrophils and reduced neutrophil extracellular traps following cerebral ischemia/reperfusion, suggesting PKM2 promotes neutrophil hyperactivation in the setting of stroke. In the filament and autologous clot/rtPA stroke models, irrespective of sex, deletion of PKM2 in myeloid cells either in wild-type or hyperlipidemic mice reduced infarcts and enhanced long-term sensorimotor recovery. Laser speckle imaging revealed improved regional cerebral blood flow in myeloid cell-specific PKM2-deficient mice that was concomitant with reduced post-ischemic cerebral thrombo-inflammation (intracerebral fibrin(ogen), platelet (CD41-positive) deposition, neutrophil infiltration, and inflammatory cytokines). Mechanistically, PKM2 regulates post-ischemic inflammation in peripheral neutrophils by promoting STAT3 phosphorylation. To enhance the translational significance, we inhibited PKM2 nuclear translocation using a small molecule and found significantly reduced neutrophil hyperactivation and improved short-term and long-term functional outcomes following stroke. Collectively, these findings identify PKM2 as a novel therapeutic target to improve brain salvage and recovery following reperfusion.