| Background:Parkinson’s disease(Parkinson’s Disease,PD)is the second largest known neurodegenerative disease in the world.At present,the known pathogenesis includes the following two aspects:Genetic factors,environmental factors,aging of nervous system and death of dopaminergic neurons caused by multiple factors.Besides,hyperphosphorylation and abnormal folding of α-synuclein,cytoplasmic clearance of α-synuclein disturbance caused by the abnormality of ubiquitin-protease pathway and autophagy-lysosomal pathway lead to the deposition of Lewy bodies.The above factors lead to the occurrence and development of Parkinson’s disease.In addition to the disturbance of cytoplasmic clearance,little is known about the extracytoplasmic clearance of α-synuclein in the pathogenesis of Parkinson’s disease and Glymphatic system as an extracellular clearance pathway may play an important role.Therefore,the study of Glymphatic system clearance dysfunction and its deep molecular mechanism in the pathogenesis of Parkinson’s Disease can provide us with a new perspective and ideas for the treatment of Parkinson’s Disease.Objective:Subacute Parkinson’s Disease(PD)model mice were established to simulate the pathological process of PD,to evaluate the ability of Glymphatic system to scavenge cytotoxic substance,and to further explore the expression,polar distribution and regulatory mechanism of aquaporin-4(AQP4),an important protein involved in Glymphatic system.We aim to explore the scavenging function of Glymphatic system mediated by AQP4 polar distribution and reveal the specific molecular mechanism of AQP4 anchoring protein α-syntrophin and its downregulation,as well as reveal the new mechanism of abnormal protein deposition and iron deposition related to the pathogenesis of PD from the point of view of Glymphatic system and to provide new intervention targets for it.Methods:8-10 weeks old C57BL/6J mice were divided into 4 groups called ABCD(Group A:Control group,Group B:MPTP group,Group C:MPTP+AAV group,Group D:MPTP+Snta1 group).Three weeks before the establishment of subacute Parkinson’s Disease(PD)model,normal saline was injected into lateral ventricle of Group Control and MPTP,adenovirus without target gene was injected into lateral ventricle of Group MPTP+AAV,and adenovirus carrying α-syntrophin gene(Sntal)was injected into lateral ventricle of Group MPTP+Snta1.After 3 weeks,the virus began to express stably.The subacute PD model was established by intraperitoneal injection of 1-methyl-4-phenyl-1(MPTP)20mg/kg/d into Group MPTP,MPTP+AAV and MPTP+Sntal for 7 days.The expressions of AQP4,and αsyntrophin in substantia nigra and striatum were detected by Western Blot method.The polar distribution of AQP-4 and the expression of α-syntrophin and NRSF proteins in nigra and striatum were observed by immunofluorescence method.Fluorescent tracer technique was used to observe and evaluate the clearance function of Glymphatic system in each group.Results:1.The function of Glymphatic system in subacute PD model mice was impaired,and the clearance rate of cerebrospinal fluid tracer decreased and accumulated in substantia nigra and striatum,while the function of Glymphatic system in subacute PD model mice could be partially restored by intracerebroventricular injection of adenovirus carrying SNTA-1 gene.At the same time,in the substantia nigra,the polarity of AQP4 protein in Group MPTP and MPTP+AAV was seriously disordered compared with that in Group Control(PB<0.05,Pc<0.001),while the polarity of AQP4 protein in Group MPTP+Snta1 was significantly better than that in Group MPTP and MPTP+AAV(PB<0.01,Pc<0.0001).In the striatum,the polarity of AQP4 protein in Group MPTP was more serious than that in Group Control(P<0.0001).The polarity of AQP4 protein in Group MPTP+Snta1 was significantly better than that in Group MPTP(P<0.01).2.Western Blot showed that the expression of AQP4 protein in striatum in Group MPTP+AAV and MPTP+Snta1 was higher than that in Group Control(PC<0.05,PD<0.01),the expression of AQP4 protein in striatum in Group MPTP+Snta1 was higher than that in group MPTP(P<0.05),and the expression of α-syntrophin protein in Group MPTP+Snta1 was higher than that in group Control(P<0.05).In the midbrain,there was no significant difference in the expression of AQP4 protein among the 4 Groups,but the expression of α-syntrophin protein in Group MPTP+Snta1 was higher than that in other 3 Groups(PA<0.01,PB,C<0.05).3.Immunohistochemical fluorescence showed that the expression of AQP4,αsyntrophin and NRSF protein in Group MPTP and MPTP+AAV was lower than that in Group Control(P<0.05),while the expression of α-syntrophin and NRSF protein in Group MPTP+Snta1 was higher than that in Group MPTP and MPTP+AAV(P<0.001).Compared with Group Control,the expression of AQP4,αsyntrophin and NRSF protein had no significant difference(P>0.05).In the striatum,the expression of AQP4 protein in Group MPTP and MPTP+Snta1 was lower than that in Group Control,the expression of AQP4 protein in Group MPTP+AAV was higher than that in Group MPTP(P<0.05),the expression of αsyntrophin protein in Group MPTP and MPTP+AAV was lower than that in Group Control(P<0.05),and the expression of NRSF protein in Group MPTP,MPTP+AAV and MPTP+Snta1 was lower than that in Group Control(P<0.001).Conclusion:1.In subacute Parkinson’s disease model mice,the polarity distribution of AQP4 in striatum and substantia nigra was disordered,and the function of Glymphatic system was impaired,which led to the decrease of its ability to scavenge harmful substances,and then deposited in striatum and substantia nigra,which played a certain role in the pathological development of Parkinson’s disease.2.NRSF-Snta1-AQP4 may be one of the pathways that affect the function of Glymphatic system.The down-regulation of NRSF protein expression in striatum and substantia nigra of subacute Parkinson’s disease model mice leads to the downregulation of α-syntrophin protein,which leads to the inability of AQP4 protein to be effectively anchored on the cell membrane,which leads to the disorder of polar distribution of AQP4 protein,and finally leads to Glymphatic system damage.3.There may be a bi-directional regulation mechanism between NRSF protein and αsyntrophin protein:the expression of α-syntrophin protein was up-regulated by intracerebroventricular injection of adenovirus carrying Snta1 gene in subacute Parkinson’s disease model mice,which could not only up-regulate the expression of AQP4 protein and improve the polarity of its distribution,but also up-regulate the expression of NRSF protein and partially restore the ability of Glymphatic system clearance. |
PDF Full Text Request