Effect Of Edaravone On Inflammatory Response And Endoplasmic Reticulum Stress After Intracerebral Hemorrhage

Objective: This study was designed to investigate the protective effect of edaravone against cerebral hemorrhage(ICH)in mice and its potential mechanism.Methods: 144 healthy male C57 BL / 6 mice(22-26g)were randomized into four groups: sham operation group(Sham group),cerebral hemorrhage group(ICH group),cerebral hemorrhage + edaravone 6 mg / kg · d(LD group),cerebral hemorrhage +edaravone 12 mg / kg · d(HD group).Each group involved more than 6 mice.Neurobehavioral tests were performed separately either before modeling or after 24 h,72h,7d,14 d modeling.Brain tissue of mice in each group were extracted to evaluate brain water content,electron microscopy,immunohistochemistry,and Western blot after 3rd day of surgery.Results: 1.Behavioral test: Sham group mice showed no obvious neurological dysfunction after surgery.Compared mice in Sham group with mice in ICH group,LD group and HD group,there were significant differences in time points(P <0.05)of tape removal time and rotating rod drop time after operation.Compared with the ICH group,both LD group and the HD group showed statistically difference(P <0.05)of tape removal time and rotating rod drop time at 3d,7d,and 14 d after operation.2.Brain tissue water content: the brain tissue water content around the hematoma of mice in the ICH group,LD group,and HD group were significantly higher than that in the Sham group at 3 days after surgery(P <0.05).The brain water content in the LD group and HD group was significantly decreased compared with that in the ICH group,which showed statistical significance(P <0.05),and the reduction of brain water content in the HD group was more significant.3.HE staining: Sham group had complete brain tissue structure,neatly arranged neurons,good morphology and structure,and its nerve cells showed no obvious edema.However,the ICH group,LD group,HD group showed inflammatory cell infiltration and the nerve cells showed vesicular degeneration.Part of the nucleus was condensed,and the chromatin density was increased.Compared with the ICH group,the pathological changes of cells in the LD group and the HD group were alleviative,which was more significantly in the HD group.4.Changes in expression of Neu N + cells: the mice n the Sham group had neatly arranged cells and good morphology cells;while in ICH group,LD group and HD group,the expression of positive cells showed decreased with nucleus shrinkage and vesicular degeneration.Compared with the ICH group,the morphological structure of LD group and HD group was significantly improved.the regions and the numbers of positive expressions were increased,which showed statistically significant(P <0.05).5.Observation of white matter fiber bundles by electron microscope: The white matter fiber bundles of the mice in the Sham group were intact and there was no obvious change in demyelination.While in the ICH group,the LD group and the HD group,we found that Myelin became significantly thinner and the demyelination occurred,and the G-ratio showed significant differences(P <0.05)compared with Sham group.The demyelinating changes in the LD and HD groups were significantly reduced compared with the that in the ICH group(P <0.05).6.The changes of CD16 /32 + cell expression: Sham group mice had slight changes of CD16 / 32 + cell expression after operation;while in the ICH group,LD group,and HD group,the expression of positive cells had significantly increased and the area of positive cells were increased.Besides,we found that the cells were branch-shaped.Compared with the ICH group,the reduction of positive expression in the LD group and HD group showed statistically significant(P <0.05).7.Inflammatory cytokine expression: The expression of IL-1β and TNF-α in the ICH group,LD group,and HD group were significantly different from those in the Sham group(P <0.05).Compared with the ICH group,the expressions of IL-1β and TNF-α in the LD group and the HD group were reduced and there were statistical differences(P <0.05).8.Immunofluorescence results: Compared with Sham group,the number of CD16 / 32 + cells and the expression of CHOP in the ICH group,LD group and HD group were increased,which showed statistical differences(P <0.05).The positive expressions were all reduced in the ICH group,LD group and HD group,which possessed statistically significant(P <0.05).9.IRE1α / TRAF2 pathway protein expression: The increased expression of Bi P,IRE1α,TRAF2 and CHOP in the ICH group,LD group,and HD group had statistical differences(P <0.05)compared with that in the Sham group.Compared with ICH group,the expressions of Bi P,IRE1α,TRAF2 and CHOP in the LD group and HD group were decreased,which were statistical significance(P <0.05).The HD group showed a more significant reduction than the LD group.Conclusion:1.After cerebral hemorrhage in mice,cell edema and apoptosis occured in nerve cells;white matter fiber bundles are injured and microglia are activated;proinflammatory factors are accumulated.2.Edaravone can reduce the edema and apoptosis of nerve cells after cerebral hemorrhage in mice and the damage of white matter fiber bundles.Meanwhile,it also can decrease the activation of M1 microglia and the accumulation of pro-inflammatory factors.3.Edaravone may inhibit the activation of M1 microglia through IRE1α / TRAF2 signaling pathway after ER stress,therefore protecting neural function.