Cdk5 Inhibitory Peptide Prevents Neurodegeneration Caused By Neurotoxicity Of P25 In Mouse Brain

BackgroundP35 is cleaved by calpain into N-terminal p]0 and C-terminal p25.Compared to p35,p25 shows longer half-life times,more soluble in cells and stronger binding to Cdk5.These differences have made p25 hyperactive Cdk5 and hyperphosphorylate the downstream substrates of the kinase,leading to neurodegenerative diseases.However,Cdk5 inhibitory peptide(CIP)which is truncated from p35,can selectively inhibit the activity of Cdk5/p25 in cultured neurons and CIP/p25 tetra-transgenic mice.In our previous studies,we demonstrated AAV9-CIP can reverse the memory loss and anxiety-like behavior observed in APP/PSI mice,reducing tau hyperphosphorylation and(Ap)deposit.In the MPTP/p induced Parkinson’s disease mouse model,we had shown that AAV9-CIP can reduce neurons loss and alleviate the motor and anxiety-like symptoms effectively.Objectives1.To establish an AAV8-GFP-p25 induced p25 overexpression mouse model.2.After injected with the AAV8-GFP-p25 virus,pathological and behavioral changes would be observed.3.Pathological changes were assessed after the treatment of AAV9-CIP.4.Eventually,reversal of behavioral abnormality afterAAV9-CIPtreatmentwould be evaluated.Methods1.We prepared the AAV8-GFP-p25(AAV8-GFP-p25)and AAV9-CIP-T2A-mCherry(AAV9-CIP)virus.2.AAV8-GFP-p25 was injected in 8 week-old mice by intracerebroventricular(i.c.v.)injection to establish the p25 overexpression mouse model,3.The model mouse was administrated with AAV9-CIP by i.c.v,injection in the same lateral ventricle one month later.4.Behavioral assays were performed in these mice at 6-month old.5.Pathological changes and the fluctuating expression of related protein between control mice,AAV8-GFP-p25 infected mice and AAV9-CIP coinfected mice were detected by immunohistochemistry and Western blot analysis.6.Statistical analysis:different types of software used were the SPSS20.0,image J and GraphPad Prism 6.05.The active avoidance response tests were analyzed by repeated-measures ANOVA,while other data were analyzed using one-way ANOVA.All values were presented as the mean of at least three determinations± SEM,and p value<0.05 was considered significant.Results1.The hyperphosphorylations of tau at multiple sites were observed in the 6 month-old AAV8-GFP-p25 mice,along with the activation of astrocytes and the elevated inflammatory factors including IL-1 and TNF-a.2.In the same time,significant anxiety-like behavior and leaning decline were detected by behavioral assays in p25 overexpression mouse model.3.Treatment with AAV9-CIP one month later after AAV8-GFP-p25 injection improved learning ability and reduced inflammatory factors in mice at 6 months old.Conclusions1.In the present study,we demonstrated that infected with AAV8-GFP-p25 in adult mice through i.c.v.injection can conduct a p25 overexpression mouse model characterized by leaning decline and anxiety-like behavior.2.It is easier to establish a p25 neuronal toxicity mouse model,saving time and money.3.Importantly,treatment of CIP after p25 neurotoxicity in adult mice would be more close to clinical scenario.4.The treatment with AAV9-CIP in p25 overexpression mice effectively reduced the pathological changes and inflammatory factors,leading to reserve learning ability.Our data concludes that CIP can protect p25 toxicity in adult mouse.