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Introduction: Microglia have a prominent role in initiating, sustaining and resolving post-stroke inflammation. The pathology of stroke largely depends on the spectrum of microglial activation with the classically (M1; pro-inflammatory) and/or alternatively activated (M2; anti-inflammatory) phenotype. It has been reported that interferon regulatory factor 5 (IRF5) and IRF4 regulate the M1 and M2 phenotype of macrophages respectively in systemic inflammation; however, it is unknown if microglial activation is regulated by the same mechanism after stroke. We hypothesize that IRF5/IRF4 directs M1/M2 microglial polarization respectively after stroke and the balance of the IRF5/IRF4 regulatory axis influences stroke outcomes.Methods: C57BL6 mice (8-12 weeks) were subjected to a 90-minute middle cerebral artery occlusion (MCAO). Stroke outcomes were evaluated at 12h, 3d, 10d of MCAO. Protein and mRNA levels of IRF4/5 at each time point were examined by IHC, Western blot and RT-PCR. Microglial phenotypes were evaluated by flow cytometry (FC) and determined by the ratio of MHC-II+ (M1) or CD206+ (M2) microglia number over the total microglia. IRF4/IRF5 conditional knockout (CKO) mice will also be utilized to examine the effect of gene deletion on stroke outcomes.Results: IRF4 protein level increased at 10d in the ipsilateral brain lysates, but IRF5 level decreased at day 10 compared to that of 3d. M2 microglia number was higher at 10d vs. 3d by FC analysis; while M1 microglia ratio was lower at 10d vs. 3d. The Iba1+IRF4+ (M2) and Iba1+IRF5+ (M1) cell number by IHC showed the same pattern as that in FC data. In addition, IRF4 mRNA in FC-sorted microglia was up-regulated at 3d of stroke and peaked at 10d, but IRF5 mRNA significantly increased at 3d vs. sham and declined to baseline at 10d. IRF5-CKO mice had significantly smaller infarct volume and improved behavior deficits than IRF5flox/flox mice while IRF4-CKO mice had worse outcomes than that of IRF4flox/flox mice.Conclusion: IRF5-mediated pro-inflammatory phenotype (M1) is predominate in the acute stage of stroke that switches to IRF4-mediated anti-inflammatory phenotype (M2) in the chronic stage. IRF5/IRF4 regulatory axis regulates the microglial M1/M2 activation and impacts stroke outcomes.