Effects And Mechanisms Of Atorvastatin On Learning And Memory Impairment Induced By Tau Phosphorylation And Neuroinflammation In Mice

ObjectiveTo investigate whether Atorvastatin（Ato）has protective effect on tau hyperphosphorylation and neuroinflammation caused by okadaic acid（OA）and its mechanism.To further clarify the neuroprotective effect of Atorvastatin on OA-mediated glial activation,learning and memory impairment,neuronal damage and its mechanisms.In order to further study the role of Atorvastatin on neurodegenerative diseases such as Alzheimer’s disease（AD）with pathological changes of tau protein,and provide theoretical basis for exploring new clinical prevention and treatment.MethodsC57BL/6 adult male mice that weighed 18-22 g were divided into five groups by random number method,mainly including the control group,OA group,Ato1 mg/kg+OA group,Ato10 mg/kg+OA group,and Ato10 mg/kg group.Mice in Ato1+OA group,Ato10+OA group,and Ato10 group were given Atorvastatin 1 mg/kg or 10 mg/kg by gavage（i.g.）daily,and equal doses of distilled water were given into control group and OA group for 6 weeks.On the 31 d of administration,the right hippocampus of each mouse was injected with OA（150 ng）or normal saline.From 36^thday,mice started Morris water maze exeriment（MWM）for six days.After MWM experiment,the morphology of vertebral neurons in the hippocampal CA1 and CA3 regions of mice were observed by using Nissl staining.Elisa was used to determinate the concentration of IL-1β、IL-6 and TNF-αin hippocampal and cortical tissue.Western blot method was used to determinate p-tau at Ser396 sites,tau5,S9sites p-GSK-3βand IL-1β,TNF-αexpression levels.The glial cell markers Iba-1and GFAP were oberved by IHC staining in the hippocampus of mice.ResultsMWM test showed that intra-hippocampal injection of OA could induced spatial learning and reference memory deficits in mice.The mice in OA group exhibited significantly higher mean escape latencies than the controls in training trial on day 3,4,and 5（P<0.01）.Compared with the OA-injected mice,the escape latencies in the mice treated with Atorvastatin（1 mg/kg or 10 mg/kg）were significantly shortened on 4^thand 5^thday（P<0.05）.In probe trial,intra-hippocampal injection of OA induced the platform quadrant entries,crossing platform times,and time in the target quadrant were reduced（P<0.05）.The swimming trajectory of the mice was chaotic,without trending.However,the OA mice treated with atorvastatin spent more time in the target quadrant and completed more platform crossing compared with OA-injected mice,the platform quadrant entries,times of crossing the platform,and the percentage of time spent in the target quadrant were increased significantly in Ato1+OA group and Ato10+OA group（P<0.05）,and the mice swimming trajectory maps had a clear trend.The result of Nissl staining showed that the number of positive neural cells in the hippocampal CA1 and CA3 region was significantly reduced in the OA group,and cells were sparsely distributed compared with those in control group.However,the hippocampal neuron loss was reduced by the treatment with Ato（1 mg/kg or 10 mg/kg）.Immunofluorescent staining showed that OA significantly upregulated the expression of Iba-1 and GFAP in all three hippocampal regions,indicating the high gliosis.Atorvastatin（10 mg/kg）treatment could effectively inhibit the upregulation of Iba1 and GFAP level.The concentration of IL-1β,IL-6 and TNF-αin hippocampus and cortex were significantly increased by ELISA,and the expression of IL-1βand TNF-αin OA group was also increased by Western blotting,which indicated that OA activated glial cells and increased the release of inflammatory factors.In the Ato10+OA group,the inflammatory factors were reduced than the OA group（P<0.01）.In addition,western blot results showed OA inhibited the expression levels of p-Akt and p-GSK-3β,increased the level of S396 p-tau（P<0.01）,on the other hand,the expression of p-Akt and p-GSK-3βincreased,the level of S396 p-tau protein decreased in the Ato10+OA group（P<0.01,P<0.05）.Conclusions1.Atorvastatin has a protective effect on the spatial learning and memory impairment induced by okadaic acid-mediated tau hyperphosphorylation and neuroinflammation in mice.2.Atorvastatin may play a protective role in okadaic acid-mediated nerve injury by up regulating Akt/GSK-3βpathway,reducing the over expression of phosphorylated tau and inhibiting the activation of glial cells.