Interferon regulatory factor 4/5 signaling impacts on microglial activation after ischemic stroke in mice

• Published in: The European journal of neuroscience Vol. 47; no. 2; pp. 140 - 149
• Main Authors: Al Mamun, Abdullah, Chauhan, Anjali, Yu, Haifu, Xu, Yan, Sharmeen, Romana, Liu, Fudong
• Format: Journal Article
• Language: English
• Published: France Wiley Subscription Services, Inc 01.01.2018; John Wiley and Sons Inc
• Subjects: Animals; Cerebral blood flow; Clinical and Translational Neuroscience; Cytokines; Enzyme-linked immunosorbent assay; Flow cytometry; Immunohistochemistry; Infarction, Middle Cerebral Artery - metabolism; Inflammation; Interferon regulatory factor; Interferon regulatory factor 4; Interferon Regulatory Factors - genetics; Interferon Regulatory Factors - metabolism; Interleukin 1; Interleukin 10; Interleukin 4; Interleukins - metabolism; Intracellular levels; IRF4; IRF5; Ischemia; ischemic stroke; Macrophages; Male; Mice; Mice, Inbred C57BL; Microglia; Microglia - metabolism; mRNA; neuroinflammation; Polymerase chain reaction; Reperfusion; RNA, Messenger - genetics; RNA, Messenger - metabolism; Rodents; Signal Transduction; Tumor necrosis factor; IRF5; IRF4; neuroinflammation; microglia; ischemic stroke
• ISSN: 0953-816X; 1460-9568; 1460-9568
• DOI: 10.1111/ejn.13778

Microglial activation is a key element in initiating and perpetuating inflammatory responses to stroke. Interferon regulatory factor 5 (IRF5) and IRF4 signaling have been found critical in mediating macrophage pro‐inflammatory (M1) and anti‐inflammatory (M2) phenotypes, respectively, in peripheral inflammation. We hypothesize that the IRF5/4 regulatory axis also mediates microglial activation after stroke. C57BL6 mice of 8–12 weeks were subject to a 90‐min middle cerebral artery occlusion, and the brains evaluated at 24 h, 3, 10 and 30 days after reperfusion. Flow cytometry was utilized to examine microglial activation and cytokine expression. RT‐PCR was performed for mRNA levels of IRF5/4 in sorted microglia. Microglial expression of IRF5/4 was examined by immunohistochemistry, and brain cytokine levels were determined by ELISA. Our results revealed that the IRF5 mRNA level in sorted microglia increased at 3 days of stroke; whereas IRF4 mRNA level exhibited biphasic increases, with a transient rise at 24 h and a peak at 10 days. The same pattern was seen in IRF5/4 protein colocalization with Iba‐1+ cells by IHC. Intracellular levels of TNF‐α and IL‐1β in microglia peaked at 3 days of stroke, and IL‐4+IL‐10+ double‐positive microglia significantly increased at day 10. Brain levels of these cytokines were consistent with microglial cytokine changes. Worse behavior test results were seen at 3 days vs. 10 days of stroke. We conclude that microglia phenotypes are dynamic to ischemic stroke, and IRF5/4 signaling may regulate microglial M1/M2 activation and impact on stroke outcomes. Our data suggest microglial M1 and M2 phenotypes are regulated by IRF5 and IRF4, respectively, in ischemic stroke. Microglial activation and IRF5/4 expression consistently exhibited a “waxed and waned” pattern in a time–course manner after stroke, providing further evidence that the IRF5 and IRF4 signaling inhibit each other along with the progression of the disease. This study represents an important new insight into the mechanism underlying the phenotypic shift of microglial activation that is related to the IRF5/4 regulatory axis in response to stroke.