Mice were subjected to 1 hour of tMCAO followed by 23 hours of reperfusion. Mice were treated with nNIF or SCR 1 hour before and 1 hour after stroke onset (10 mg/kg). Open circles, females; filled circles, males. (A) Brain sections were stained with TTC. Red areas indicate healthy brain tissue; white areas show infarcted brain tissue (outlined with black dotted line). (B) Quantification of brain infarct volumes 24 hours after stroke. (C) Bederson’s test was used to assess neurological outcome 24 hours after stroke. (D) Twenty-four hours after stroke, motor function was measured using the grip test. n = 14 per group. (E) In a separate experiment, mice were followed for up to 7 days after stroke; survival was monitored. n = 9 for SCR; n = 10 for nNIF. (F–H) NETs in brain tissue were identified by staining for MPO (red), H3cit (green), and DNA (DAPI; blue). The percentage of NET-forming neutrophils was quantified by counting of H3cit⁺ neutrophils. n = 5 per group. Scale bar: 50 μm. (I) Plasma NETs were measured 24 hours after stroke using MPO-DNA complex ELISA. n = 7–9 per group. (J–L) Brain sections were stained for apoptosis by labeling of DNA strand breaks with TUNEL (green). TUNEL⁺ cells were counted in the striatum and cortex. n = 5 per group. Scale bar: 100 μm. (M–O) Flow cytometric analysis of single-cell suspensions of ipsilesional brain hemispheres. CD45⁺CD11b⁺Ly6G⁺ cells were counted in nNIF- and SCR-treated animals. n = 5–7 per group. (P) Quantification of neutrophil staining with Ly6G in brain sections 24 hours after stroke. n = 5 per group. Groups were compared by unpaired t test (B, H, I, O, and P), Mann-Whitney test (C and D), ordinary 1-way ANOVA (L), or log-rank test (E). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.