The Role And Mechanism Of APOE4-mediated Microglial Phagocytosis In Exacerbating White Matter Damage After Experimental Subarachnoid Hemorrhage

Objective: Apolipoprotein E(apoE,protein;APOE,gene)is closely related to the process of white matter injury.Although the role of APOE genotype in acute brain injury is beginning to receive attention,the specific mechanism remains unclear.Our aim was to further investigate the possible mechanism of APOE genotype in early white matter injury(WMI)after subarachnoid hemorrhage(SAH).Methods: The endovascular perforation model of SAH or sham surgery was performed in a total of 177 male C57BL/6 mice(weight 22-26g).The adeno-associated virus(AAV)was used to express targeted microglial AOPE in mice and lentivirus was used to express AOPE in BV2 cells,respectively.To find the possible reasons of difference between APOE3-BV2 cells and APOE4-BV2 cells in bleeding environment,the high throughput sequencing was used.Neurobehavioral tests,immunohistochemistry,Western Blotting,RT-q PCR,flow-cytometric studies,transmission electron microscopy,reactive oxygen species(ROS)and JC-1 test were performed.Results: Endogenous myelin expression was decreased and reached the bottom at 48 h after SAH.apo E was increased after SAH and co-located with microglia in the region of white matter.APOE4 significantly decreased myelin level and decreased neurobehavioral function.Further tests of microglia phagocytosis in mice showed that APOE4 induced increased debris myelin,increased CD86 and decreased CD206.Indexes of phagocytosis showed that the phagocytosis of APOE4-BV2 microglia on latex beads decreased,Arg-1decreased,CD86 and CD16 increased,suggesting that the phagocytosis function of BV2 microglia was weakened after oxy Hb.High-throughput sequencing revealed that APOE polymorphism could affect autophagy,oxidative stress and other functions.The following experiments proved APOE4 increased ROS,decreased JC-1 poly to monomer ratio,and increased DHE content in white microglia or matter region.Meanwhile,APOE4 made the mitochondria of microglia smaller and rounder.Conclusion: Our finding demonstrated that APOE4 significantly aggravated the WMI and decreased neurobehavioral function.This more severe WMI and poorer neurobehavioral function may be due to microglial phagocytosis after SAH damage.Further mechanism exploration found that the damaging effects of APOE4 in SAH may be associated with microglial oxidative stress-dependent mitochondrial damage.Inhibiting mitochondrial oxidative stresscan enhance the phagocytic function of microglia.In conclusion,anti-oxidative stress and phagocytosis protection may serve as promising treatments in the management of SAH.Therefore,anti-oxidative stress and phagocytosis protection targeting APOE polymorphism may serve as promising treatments in the management of SAH patients.