Background: Neutrophils are the initial peripheral immune cells to be recruited following ischemic stroke, playing a dual role in tissue repair and injury. Elevated peripheral neutrophil counts correlate with larger infarct volumes and poorer outcomes, particularly in elderly patients. Aging alters neutrophil function, including increased expression of the nuclear transcription factor NR4A2, a regulator of cell survival and inflammation. We sought to investigate the role of NR4A2 in neutrophil recruitment, persistence, and function in aged brains following ischemic stroke using a murine transient middle cerebral artery occlusion (tMCAO) model. Methods: Young and aged Catchup (Ly6g-cre with a tdTomato reporter) mice were crossed with Nr4a2 floxed mice to generate Catchup Nr4a2-/- mice and were subjected to tMCAO for 90 minutes. Neutrophil infiltration and persistence were tracked through pulse labeling with 5-ethynyl-2’-deoxyuridine (EdU). Neutrophil distribution was quantified on coronal sections through wide-field immunofluorescence imaging at various timepoints. Results: Neutrophil distribution post-stroke was markedly altered in aged mice, with a higher concentration near the cortical surface at 72 hours (Fig. 1A and B), despite similar total counts. EdU labeling revealed that neutrophils infiltrated the brain within the first 24 hours and persisted for up to 120 hours post-stroke. EdU-positive cells replicated the spatial distribution of total neutrophils, confirming prolonged survival. Preliminary functional assays showed reduced metabolic activity in aged neutrophils, consistent with cellular senescence (Fig. 1C). Conclusion: Our findings demonstrate that aging alters neutrophil infiltration patterns and prolongs their survival in the infarcted brain, contributing to sustained inflammation and secondary injury. NR4A2 plays a critical role in regulating these processes, particularly in aged neutrophils. Future studies will explore the feasibility of targeting NR4A2 as a novel therapeutic target to mitigate age-related secondary injury.
Sullivan et al. (Thu,) studied this question.