The Role Of DISC1 Aggregates-positive Microglia In Aging-related Cognitive Decline

Objective:Microglia are macrophages and immune cells in the central nervous system,which play pivotal roles in neurodevelopment and brain aging.Exploration the molecular mechanism underlying the maintainence of microglial homeostasis would hopefully bring novel targets for neurological disorders related to both neurodevelopment and aging.Disrupted-inSchizophrenia-1(DISC1)is a genetic susceptibility gene for a variety of psychiatric diseases,including schizophrenia.Dysfunction of DISC1 signaling contributes to various neurodevelopmental disorders such as autism spectrum disorders(ASD),neuropsychiatric disorders such as schizophrenia,biopolar and Alzheimer’s disease(AD).In contrast to the roles of DISC1 in neurons and neural stem cells,the roles of DISC1 in microglia remain unknown.We have observed that DISC1 is expressed on microglia and aggregated on some microglia.This thesis analyzes the spatiotemporal expression profiles of DISC1 aggregatespositive microglia during brain development and aging.We also preliminarily explore the function of endogenous DISC1 and DISC1 aggregates in regulating microglia function and its effect on brain cognition.Methods:(1)The expression of DISC1 in microglia was confirmed by real-time quantitative PCR,western blotting and immunofluorescence staining.The proportion of DISC1 aggregates-positive microglia in the hippocampus at different ages was analyzed to construct the spatiotemporal expression profile of DISC1.(2)The relationship between DISC1 aggregates-positive microglia and cellular senescence in the brain of aging mice and AD model mice was analyzed by immunofluorescence staining.BV2 cells and primary cultured cortical microglia were treated with LPS to construct the senescent cell model to analyze the protein level of insoluble DISC1 and the formation of DISC1 aggregates.The phagocytosis of DISC1 aggregates-positive microglia was further analyzed by observing the phagocytosis of latex beads by microglia on acute mouse brain slices.(3)After knockdown of Disc1 by siRNA on BV2 cells,the phagocytosis and inflammatory response of BV2 cells were analyzed.Disc1fl/fl mice and Cx3cr1-CreERT2 tool mice were used to construct conditional knockout mice with specific knockout of Disc1 on microglia.The behavioral performance of mice was analyzed by open field,elevated plus maze,new object recognition and three box social test.Results:1.DISC1 is expressed in mouse microglia and aggreagetd in some microglia.The proportion of DISC1 aggregates positive microglia showed dynamic changes during brain development and aging.The expression of Disc1 mRNA and DISC1 protein was detected in the murine microglia cell line BV2 and in microglia which were derived from primary cortical cultures of neonatal mice.In addition,DISC1 was found to be aggregated around the nucleus of some microglia.The proportion of DISC1 aggregates-positive microglia exhibited dynamic change during the development and aging of hippocampus in C57 mice.The increasement occurred in P0-P15 stage and 7-25-month old stage respectively,and tended to increase with the further aging of mice.2.DISC1 aggregates-positive microglia are senescent microglia.DISC1 aggregates-positive microglia are senescent microglia in the brain of aged WT mice and AD mice.The numbers of DISC1 aggregates-positive microglia were increased by AD environment.DISC1 aggregates positive microglia existed in the region far away from Aβ in the brain of APP/PS1 mice.DISC1 aggregates-positive microglia habour impaired phagocytosis.3.Cellular senescence leads to the increase of DISC1 insoluble fraction and promotes the formation of aggregates.Induction of senescence by LPS treatment exacerbates insolubility of DISC1 and formation of a aggregates in microglia.It is suggested that DISC1 aggregates+microglia will be dysfunctional due to the reduction of soluble DISC1.4.Deficiency of DISC1 in microglia impairs microglial homeostasis,leading to cognitive deficits in mice.Knocking-down of Disc1 led to impaired phagocytosis and excessive inflammatory response of BV2 cells.Conditional knock out of Disc1 in microglia resulted in decreased spontanouse locomotor activity,anxiety,memory deficits in recoganizing novel object and social interaction in mice.Conclusion:DISC1 aggregates-positive microglia exhibited two proportional increases in neurodevelopment(P0-P15)and aging respectively.In aging brain,DISC1 aggregates positive microglia are senescent cells and exhibiting impaired phagocytosis.Both aging and AD environment enhances the numbers of DISC1 aggregates positive microglia.Such DISC1 aggregates in microglia is likely due to the accumulation of insoluble DISC1.DISC1 aggregates were formed by insoluble DISC1,which was exacerbated by cellular senescence induced by LPS.Deficiency of DISC1 results in impaired phagocytosis,while enhanced inflammatory response of BV2 cells.Abalation of DISC1 in adult microglia results in cognitive deficits in mice.In summary,the current results of this thesis suggest that the increase of insolubility of DISC1 on microglia led to the formation of aggresomes during aging,which in turn leads to the decrease of endogenous soluble DISC1 level and the loss of DISC1 function in microglia.The functional loss of DISC1 will lead to the senescent characteristics of microglia,which will lead to cognitive impairment in mice.