Microglia In Hemorrhagic Brain Injury:Mechanisms And Clinical Implications

Background and Objective:Intracerebral hemorrhage(ICH)is a devastating disease associated with high mortality and disability,but no effective treatment exists.After ICH onset,immune and inflammatory responses contribute to the disruption of blood brain barrier(BBB),formation of edema that surrounds hematomas,and the cell death process,resulting in aggravated brain injury.As resident immune cells in the brain,microglia are the first responders to ICH.Following ICH,microglia acquire properties of phagocytosis,antigen presentation,and the production of immune factors such as the proinflammatory cytokines,tumor necrosis factor-?(TNF-?),interleukin-1 beta(IL-1?),and the anti-inflammatory cytokines IL-10,transforming growth factor-?(TGF-?).However,the precise role of microglia in ICH injury remains unclear.The survival of microglia depends on colony stimulating factor 1receptor(CSF1R)signaling.In this study,we determined the effects of a CSF1R inhibitor(PLX3397)on microglia and ICH injury using two mouse models of ICH.18-kDa translocator protein(TSPO)is a commonly used marker of actvated microglia.Then we used a TSPO ligand extifoxine to determine the impact of pharmacological modulation of microglia on ICH injury.Methods:To deplete microglia,mice received daily treatment with vehicle PBS or PLX3397(40 mg/kg)by oral gavage for 21 days prior to ICH induction by injection of autologous blood or collagenase.Etifoxine or saline vehicle was administrated at50 mg/kg by intraperitoneal injection immediately.Neurodeficits,brain hematoma and edema volumes were measured to to assess the therapeutic effecacy of Etifoxine.Modified Neurological Severity Score(mNSS)and corner turning test were used to evaluate neurodeficits.Magnetic resonance imaging(MRI)and dry-wet weight measurements were performed to assess the volume of hematoma and the brain edema.Flow cytometry and ELISA assays were done to measure immune cells in the brain and periphery.BBB disruption was measured using MRI,immunostaining and western blot.Results:Microglia was activated and produced amounts of inflammation mediators after ICH in humans and mice.(1)CSF1R inhibitor PLX3397 effectively depleted microglia,and the depletion of microglia was sustained after ICH.Importantly,CSF1R inhibition attenuated neurodeficits and brain edema in two experimental models of ICH induced by injection of collagenase or autologous blood.The benefit of CSF1R inhibition was associated with reduced leukocyte infiltration in the brain and improved BBB integrity after ICH.(2)TSPO was up-regulated i microglial cells from brains of patients with ICH and in CD45~intnt CD11b~+microglial cells from mice subjected to collagenase-induced ICH.And the exogenous ligands of TSPO,Etifoxine significantly reduced the activation of microglia and microglial production of proinflammatory cytokines IL-6 and TNF-a,and the subsequent infiltration of immune cells,furthermore improved BBB integrity and diminished cell death.Notably,the protective effect of Etifoxine was abolished in mice depleted of microglia by using the CSF1R inhibitor PLX3397.Conclusions:Our results suggest a detrimental role of microglia in ICH injury.Pharmacological modulation of TSPO may attenuate ICH injury and improve disease outcome.