| Objective: Use dementia rat model was injected with ibotenic acid(IBO)into bilateral Meynert basilar nucleus and autoimmune thrombus-induced multiple cerebral infarction(MID)of dementia rat model.To investigate the effect of EI injection on learning and memory function of dementia rat model,the influence of brain energy charge,brain pathomorphology,insulin content and PI3K/AKT signaling pathway-related protein expression,and to explore the effect of EI injection against dementia and PI3K/AKT signal pathway regulates the relationship between brain energy and preliminary reveals the molecular mechanism of antidementia by EI injection.Methods:1.EI injection safety measurementKM mice were randomly divided into EI injection male group,EI injection female group,solvent male group and solvent female group according to gender and body weight.Each group was given intravenous injection of the corresponding drug,observed acute EI injection on mice Toxic reaction.SD rats were randomly divided into EI high dose(2mg/kg),EI low dose group(1mg/kg),solvent group,and blank group according to body weight and exercise time.Each group was given intravenous injection of the corresponding drug,continuous 8 days;The behaviors of rats in each group were tested by climbing bar test,opening test and Morris water maze test;Blood tests were performed on rats in each group using a fully automated hematology analyzer;Blood biochemical tests were performed on each group of rats using a fully automatic biochemical analyzer.2.Study on effect and mechanism of EI Injection Anti-dementia2.1.Model establishment and intervention:The rat model of dementia was established by bilateral injection of IBO into the basal ganglia of Meynert.The rat model of MID dementia was established by autologous thrombus injection.After successful rat model establishment,IBO dementia model rats were divided into model group,donepezil hydrochloride tables group,EI high dose group(2mg/kg),EI low dose group(1mg/kg)and sham operation group.The MID model rats were divided into model group,donepezil hydrochloride tables group,co-dergocrine mesylate tables group,EI high dose group(2mg/kg),EI low dose group(1mg/kg)and sham operation group.The EI injection was intravenously injected at 2 ml/kg once daily for 8 weeks.2.2 pharmacodynamic study: Morris water maze test was used to investigate the effect of EI injection on the behavior of the two models of rats.The effect of EI injection on the learning and memory function of dementia rats was evaluated.The effects of EI injection on ?-amyloid protein deposition and pyramidal cells in the brains of two rat models were investigated by Congo red staining.2.3.the study of brain energy charge: The contents of ATP,ADP and AMP in the brains of two rat models of EI injection were detected by HPLC,and the brain energy charge was calculated.The effect of EI injection on brain energy charge of dementia rat model was evaluated.2.4 study of PI3K/AKT signaling pathway: ELISA was used to detect the effect of EI injection on insulin content in brain tissue of rats.Western blot was used to detect the effect of EI injection on the expression of p-PI3 K and p-AKT protein in the brains of rats.Results:1.EI injection safety evaluationThe results of acute toxicity test in mice showed that after administration of EI injection,the spirit,hair and movement status of mice in each group were normal within 14 days.There was no significant influence on body weight and food consumption of mice in each group(P>0.05),and no obvious adverse reactions.None of the mice died.The results of normal rat behavioral experiments showed that the climbing experiment showed no significant difference in scores between groups,and the results of the opening experiment showed that the exercise time of rats in the high-dose EI group was significantly lower than that of the sham-operated group(P<0.05).There was no significant difference between the exercise time and exercise distance in other groups(P>0.05).Morris water maze results showed that the time to enter the quadrant quadrant was significantly prolonged in the space exploration of EA low dose group compared with solvent group rats(P< 0.05)There was no significant difference in the number of escape latency,time to enter the quadrant quadrant and platform quadrant of the other training groups in the 5th day(P>0.05).The blood test results of normal rats showed that the blood routine indicators and blood biochemical indexes in each group were also No significant difference(P>0.05).2.Effects of EI injection on learning and memory function in model rats2.1Effect of EI injection on learning and memory function in IBO model rats with dementiaThe Morris water maze directed navigation test results showed that compared with sham operation,the escape latency of the model group rats on the 1st,2nd,3rd,and 5th days of the training was significantly prolonged(P<0.01),and the 2nd dose of the EI injection 2 mg/kg dose group was the 2nd day.The fifth-day escape latency was significantly shortened(P<0.01).The escape latency of EI injection low dose group was significantly shorter on the second,fourth,and fifth days(P<0.01).The results of space exploration experiments showed that the model group rats Compared with the sham-operated group,the number of crossing platform quadrants,number of platform crossings,and platform crossing times decreased significantly(P<0.05).Compared with the model group,the rats of the EI injection high and low groups crossed the platform.The quadrant time was significantly increased(P<0.05);although the number of crossing platforms and the number of platform crossings in each dose group of EI injection increased,there was no statistical significance(P>0.05).2.2 Effect of EI injection on learning and memory function in MID model ratsThe results of Morris water maze orientation route indicated that compared with sham operation,the escape latency of the model group rats on the 1st,3rd,and 5th day of training was significantly prolonged(P<0.01),and the EI injection high,low dose group on the 1st The 3d and 5d escape latency was significantly shortened(P<0.05).The results of space exploration experiments showed that compared with the sham operation group,the rats in the model group significantly reduced the number of platform crossings and the number of platform crossings(P<0.05).Compared with the control group,the number of rats passing through the platform and the number of platform crossings in the EI injection high dose group increased significantly(P<0.05).Although the EI injection dose group quadrant retention time and crossing platform quadrant times increased,However,it was not statistically significant.3.Effect of EI injection on ?-amyloid protein deposition and pyramidal cells in two dementia model rats3.1Effects of EI Injection on ?-amyloid protein deposition and pyramidal cell in IBO dementia model ratsThe results of Congo red staining showed that the hippocampal CA1 and cortical areas of the model group stained deeper than the sham group,and they were widely distributed.There were amyloid deposits,scattered distribution,pyramidal cell pyknosis,disordered arrangement and loosening;the EI injection was high.A small amount of amyloid deposits were found in the CA1 and cortical areas of the low-dose group.The staining results were shallow,the layout was limited,and the pyramidal cells were arranged more neatly.3.2 Effect of EI Injection on ?-amyloid protein deposition and pyramidal cell in MID dementia model ratsThe results of Congo red staining showed that the hippocampal CAI and cortical areas of the model group had darker staining,scattered amyloid protein distribution,pyramidal cell shrinkage,and looser arrangement than the sham operation group;EI injection high and low dose group CA1.The area and cortical areas became lighter,showing a small amount of amyloid deposition,less distribution,and pyramidal cells arranged more neatly.4.Effect of EI injection on brain energy charge in rat model4.1 Effect of EI Injection on brain energy charge in IBO dementia ratsCompared with the sham operation group,the ATP and ADP contents in the brain tissue of the model group decreased(P>0.05),the AMP content increased(P>0.05),and the energy load decreased(P>0.05).Compared with the model control group,the contents of ATP and ADP in the brains of rats with high-and low-dose EI injections increased(P>0.05),the AMP content decreased(P>0.05),and the energy load increased.(P>0.05),suggesting that EI injection has no significant effect on the brain energy charge of this model.4.2 Effect of EI Injection on brain energy charge in MID model ratsCompared with the sham-operated group,the contents of ATP and ADP in the brain tissue of the model group were significantly lower(P<0.05),the AMP content was significantly increased(P<0.01),and the energy load was significantly reduced(P<0.05);In comparison,the contents of ATP and ADP in the brain of rats with high dose of EI injection were significantly higher(P<0.05),and the content of AMP was significantly lower(P<0.05).The ATP content in the brain of rats in the low-dose EI injection group was significant.Increased(P<0.05),ADP increased(P>0.05),AMP decreased(P>0.05),but it was not statistically significant(P>0.05);EI injection in rats brain tissue Energy load increased significantly(P<0.01).5.Effects of EI Injection on related indexes of PI3K/AKT signaling pathway in two dementia model rats5.1 Effects of EI Injection on related indexes of PI3K/AKT signaling pathway in IBO dementia ratsCompared with the sham-operated group,the insulin level in the brain of the model group was significantly lower(P<0.05).Compared with the model group,the insulin level in the brain of the high-dose EI group was significantly increased(P<0.05).The low-dose injection group showed a trend of increasing,but it was not statistically significant(P>0.05).Compared with the sham-operated group,the model group had a decreasing trend of pPI3K/PI3 K and p-AKT/AKT(P>0.05);EI injection Compared with the model group,the levels of p-PI3K/PI3 K and p-AKT/AKT in the fluid group increased,but there was no statistical significance(P>0.05).5.2 Effects of EI Injection on related indexes of PI3K/AKT signaling pathway in MID Model RatsCompared with the sham-operated group,the insulin level in the brain of the model group was significantly decreased(P<0.01).Compared with the model group,the insulin level in the brain of the high-dose EI group was significantly increased(P<0.05).The EI injection in the low-dose group had a tendency to increase,but it was not statistically significant(P>0.05).Compared with the sham-operated group,the p-PI3K/PI3 K,pAKT/AKT in the model group was significantly lower(P<0.05);Compared with the model group,p-PI3K/PI3 K was significantly increased(P<0.05),and p-AKT/AKT was increased in the low dose group(P<0.05).Conclusion:The safety of EI injection injection was good;EI injection could improve the learning and memory function of two rat models of dementia,had a good antagonistic effect on brain ?-amyloid deposition,and could protect nerve cell injury;EI injection IBO-induced dementia model rat brain energy charge has no significant effect;but can significantly increase the level of brain energy charge in MID model rats.The regulation of brain energy load in this model is closely related to PI3K/AKT signaling pathway. |
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