The Role Of Microglial Phenotype In Epileptogenesis

PART? THE DYNAMIC ALTERATION OF MICROGLIAL PHENOTYPEDRUING THE PROCESS OFEPILEPTOGENESISObjective: To explore the dynamic alteration of microglial phenotype and associated cytokines druing the process of epileptogenesis.Methods: 7-8 weeks old C57BL/6J mice were randomly divided into treatment group and control group.The mice in the treatment group were induced status epilepticus(SE)by pilocarpine intraperitoneally,and were monitored by video and EEG in the meantime.The changing trend of the proportion of microglial phenotypes M1 and M2 were detected by flow cytometry;and the concentration of the phenotype associated cytokines IL-1??IL-4 and IL-10 were detected by enzyme-linked immuno sorbent assay(ELISA).Results:(1)Pilocarpine administration scussessfully induced both SE behavior and nerve electrophysiology performance in C57BL/6J mice.The EEG records after pilocarpine administraton were consistent with different stages of epileptogenesis.(2)As long as SE was induced the proportion of microglia underwentvariations throughout the stages of epileptogenesis.The percentage of M1(CD16/32+ cells)increased to a peak level within the 1st day following SE,subsequently maintained at this high level and then gradually reduced back to the normal control level from day 20 to day 28 after SE.In contrast,the percentage of M2(CD206+ cell)exhibited an immediate drop on the 1st day after SE and then began to gradually increase until reaching a peak at 20 th day after SE.Subsequently,the percentage of M2 then decreased to control levelsduring the fourth week after SE.(3)The altered trends in the microglia-associated cytokines IL-1b(M1),IL-4and IL-10(M2)paralleled the changes in phenotype proportions.Conclusion: The proportion of microglia underwentvariations throughout the stages of epileptogenesis.The altered trends in the microglia-associated cytokines IL-1?(M1),IL-4and IL-10(M2)paralleled the changes in phenotype proportions.Indicating that the changing of microglial phenotype are involved in the process of epileptogenesis.PART? INTRAPERITONEAL INJECTION OF IFN-?/IL-4 MODULATES THE PHENOTYPE OF MICROGLIAAND ASSOSIATED CYTOKINES IN BRAINObjective: Toexplore wehther intraperitoneally administrating IL-4 and IFN-?could modulate the phenotype of microglia in brain.Methods: According to the result of part ?,M1 proportion increased significantly in the early stage after SE while M2 proportion meet a sharp increase in the middle stage after SE.In order to explore the method of regulating microglia,mice were treated by IFN-? and IL-4to further promote the increase of M1 proportion and inhibit the increase of M1 proportion respectively in the early stage after SE;the same treatment was undertook to inhibit the production of M2 and further promote the increase of M2 proportion in the middle stage after SE.The alteration of the proportion of M1 and M2 were detected by FCM;and the concentration of the phenotype associated cytokines IL-1 ?? IL-4 and IL-10 were detected by ELISA.Results:(1)Intraperitoneal injection of cytokine is able to increase the concentration of the cytokine in peripheral serum and brain tissue.(2)Early supplementation with IFN-? did not further increase the proportion of M1 after SE.Supplement of IL-4 in the middle stage can not further increase the proportion of M2 neither.The peak concentration of IL-1,IL-4 and IL-10 in these two groups ware not significantly different from that of control group.(3)Early administration of IL-4 significantly inhibited the increase of M1 in early stage after SE.Similarly,the administration of IFN-?inhibited the elevation of M2.The peak concentration of IL-1,IL-4 and IL-10 in these two groups was significantly decreased than that of saline group.(4)Inhibition of phenotypic fluctuations is not achieved by inhibition of microglial activation.Conclusion: Treatment to further increase the degree of phenotypic fluctuation of microglial phnetype after SE did not achieve as expect,but the treatment to inhibit the fluctuation of microglial phenotype after SE got the ideal ending.PART ? THE EFFECT OF INHIBITATION OF MICROGLIAL PHENOTYPE FLUCTUATION AFTER SEON THE OUTCOME OF EPILEPSY AND CONGNITIVEIMPAIRMENTObjective: To explore the effect of the inhibition of microglial phenotypefluctuationafter SE on the outcome of epilepsy and cognitive impairment.Methods:Mice in experimental group(SE group,SE-IL-4 group,SE-IFN-? group)and control group were implanted with intracranial electrodes 50 days after SE and continuous EEG detection for 14 days.The duration,frequency and sevierty of SRS were analysed.Morris water maze was conducted to determine the spatial learning and memory ability of mice.Results:In the process of EEG detection,the control group(normal control group,IFN-? control group,IL-4 control group)did not appear seizures.There was no significant difference in the incidence of SRS in SE-IL-4 group,SE-IFN-? group,and the SE group,but the duration,frequency and severity of SRS in SE-IL-4 group and SE-IFN-? group were significantly better than those of the SE group.In SE-IL-4 group and SE-IFN-? group,the Morris water maze performance of was better than SE group,but still can not catch up with the normal control group.Conclusion: According to the microglial phenotypic variation tendency after SE,precisly inhibiting the phenotype fluctuation can improve the outcome of epilepsy,reduce the duration,frequency and severity of SRS,and ameliorate the damage of cognitive function after SE.