Transforming Growth Factor Beta-Activated Kinase 1–Dependent Microglial and Macrophage Responses Aggravate Long-Term Outcomes After Ischemic Stroke

Abstract

Background and Purpose— Microglia/macrophages (Mi/MΦ) can profoundly influence stroke outcomes by acquiring functionally dominant phenotypes (proinflammatory or anti-inflammatory; deleterious or salutary). Identification of the molecular mechanisms that dictate the functional status of Mi/MΦ after brain ischemia/reperfusion may reveal novel therapeutic targets for stroke. We hypothesized that activation of TAK1 (transforming growth factor beta-activated kinase 1), a key MAP3K upstream of multiple inflammation-regulating pathways, drives Mi/MΦ toward a proinflammatory phenotype and potentiates ischemia/reperfusion brain injury. Methods— Young adult mice were subjected to 1 hour of middle cerebral artery occlusion (MCAO) followed by reperfusion. TAK1 was targeted by tamoxifen-induced Mi/MΦ-specific knockout or administration of a selective inhibitor 5Z-7-Oxozeaenol after MCAO. Neurobehavioral deficits and long-term gray matter and white matter injury were assessed up to 35 days after MCAO. Mi/MΦ functional status and brain inflammatory profiles were assessed 3 days after MCAO by RNA-seq, flow cytometry, and immunohistochemistry. Results— TAK1 Mi/MΦ-specific knockout markedly ameliorated neurological deficits in the rotarod and cylinder tests for at least 35 days after MCAO. Mechanistically, RNA-seq of purified brain Mi/MΦ demonstrated that proinflammatory genes and their predicted biological functions were downregulated or inhibited in microglia and macrophages from TAK1 Mi/MΦ-specific knockout mice versus WT mice 3 days after MCAO. Consistent with the anti-inflammatory phenotype of Mi/MΦ-specific knockout, oxozeaenol treatment mitigated neuroinflammation 3 days after MCAO, manifested by less Iba1 + /CD16 + proinflammatory Mi/MΦ and suppressed brain invasion of various peripheral immune cells. Oxozeaenol treatment beginning 2 hours after MCAO improved long-term sensorimotor and cognitive functions in the foot fault, rotarod, and water maze tests. Furthermore, Oxozeaenol promoted both gray matter and white matter integrity 35 days after MCAO. Conclusions— TAK1 promotes ischemia/reperfusion-induced inflammation, brain injury, and maladaptive behavior by enhancing proinflammatory and deleterious Mi/MΦ responses. Therefore, TAK1 inhibition is a promising therapy to improve long-term stroke outcomes.