Effect And Mechanism Research Of Guben-Jiannao In Rat With AD

ObjectivesWith the rapidly aging population, Alzheimer’s Disease (AD) is becoming more and more widely attention, because of its high prevalence, morbidity and impact on the society and their families.Therefore, the whole world was aware of strengthening research AD’s etiology and pathogenesis, and the development of the clinical value of effective control measures and the drugs and the practical significance on this basis. Our study is going to discuss on treatment and mechanism of AD by Guben-Jiannao method. Based on this, we can interpret the scientific connotation of the theory on Guben-Jiannao method, seek for the Chinese medicine to improve learning, memory and clinical treatment of AD, rich up the contents of TCM theories on Guben, prevention and treatment of AD.Methods1.Theoretical Discussion:Using the method of literature research, a systematic review of traditional Chinese medicine theory of AD from the essence of life and kidney-spleen. Combining with the relevant research results of the team, we systematically elaborated up the rule and treatment theory on Guben-Jiannao.2.Experimental Study:Around excessive phosphorylated tau protein is the core mechanism of Alzheimer’s disease, In order to curbing excessive tau protein phosphorylation to protecting dendritic spines for the targets, choosing Aricept as the western medicine control, used the Guben-Jiannao method of intervention on AD model rats, we observed its effects on the AD model and explored mechanism of its action.We bought 70 clean Wistar rats aged of 12 months, half and half for male and female. We divided them randomly into 7 groups after adaptive feed for 7 days. Such as the normal group, blank control group, model group, Guben-Jiannao fluid high dose group (short for high group), middle group, low group and Aricept group,10 in each group. Normal group would not be processing.Model group and treatment groups were given injection concentration of 5μg/μl A β1-42 in bilateral hippocampal brain stereotaxic to copy AD model, blank control group rats were given physiological saline injections by the same place and dose. After 3 days, each treatment group had been given corresponding drug lavage by respectively according to the 10 ml/kg. d for 4 weeks. Blank control group and model group had been given saline lavage in accordance with the same dose for 4 weeks.After lavage, each group rats were put into Morris Water Maze (MWM) for navigation experiments and space exploration experiment, which was to be observed up the influence of the behavior of AD rats treated by Guben-Jiannao method. And also, we used Golgi staining to observe rats’hippocampal neuron dendritic spines shape change and density change, used RT-PCR method to detect expression level of EphB2 and PARP 1, used Western Blot method to detect expression of total tau, P-tau (Ser199 Ser396, Thr231 loci), EphB2 and PARP-1. Based on these, we discussed Guben-Jiannao method’s effects on the AD model and explored mechanism of its action.Results1. Influence of behavioral science on AD rat model by Guben-Jiannao methodIn navigation experiment, The incubation period of the model group rats to find hidden underwater platform was longer than normal group (P<0.01), the incubation period of treatment groups were shorter than the model group (P<0.01), the incubation period of high group and middle group were shorter than the Aricept group (P <0.01); Percentage comparison of target quadrant swimming time of model group significantly lower than normal group (P<0.01), the treatment groups were obviously higher than that of model group (P <0.01), and middle group was higher than Aricept group (P< 0.05). In space exploration experiment, the model group rats to find the original sets of the incubation period significantly longer than the normal group (P<0.01), the incubation period in treatment group were shorter than the model group (P<0.01), and high fixation group and solid in the incubation period is shorter than the Aricept group (P<0.05); Model group rats through the original position significantly less than normal group (P<0.01), the incubation period in treatment group were more than model group (P<0.01), and high fixation group and solid than Aricept group (P<0.01). To compare with normal group, Target quadrant swimming time percentage of model group was significantly lower (P<0.01), that of the treatment group were obviously higher than that of model group (P <0.01), high group and middle group are both higher than that of Aricept group (P<0.05). These results showed that the research of AD model was successful and Guben-Jiannao method obviously improved AD rats’learning and memory functions.2. Influence on dendritic spines in Hippocampal neurons of AD rat model by Guben-Jiannao methodDensity of Hippocampus-neuron-dendritic-spines in Model group rats is significantly lower than normal group (P<0.01), that in treatment groups were obviously higher than that of model group (P <0.01), that in high group and middle group were higher than Aricept group (P<0.01). These results showed up that Guben-Jiannao method could obviously improve the hippocampal neuron dendritic spines morphology of AD rat.3. Influence on P-tau on AD rat model by Guben-Jiannao methodWe detected P-tau by Western blot, which showed that P-tau in the model group increased significantly than that in the normal group (P<0.01), the treatment group significantly lower than the model group (P<0.01), high group and middle group were both lower than Aricept group (P<0.01).4. Influence on expression of EphB2 on AD rat model by Guben-Jiannao methodWestern blot and RT-PCR method had the same results. EphB2 expression in model group significantly lower than normal group (P <0.01), that in each treatment group was significantly higher than model group (P<0.01), high group and middle group were both higher than that in Aricept group (P<0.05).5.Influence on expression of PARP-1 on AD rat model by Guben-Jiannao methodWestern blot and RT-PCR method also had the same results. PARP-1 expression in model group significantly higher than normal group (P<0.01), that in each treatment group was significantly lower than model group (P<0.01), and high group was lower than that in Aricept group (P<0.05).Conclusions1. The onset of AD closely related to spleen and kidney. Deficiency of spleen and kidney, lack of pulp, both phlegm and blood stasis are the basic pathogenesis. Guben-Jiannao method is an effective therapy for treatment of AD.2. Guben-Jiannao method can obviously improve the learning and memory ability of AD model rats, and reduce the hippocampus pathomorphism changes; increase expression of EphB2, reduce PARP 1 level and inhibit excessive phosphorylation of tau protein.3. Guben-Jiannao method can effectively prevent and treat AD and improve its clinical symptoms. The mechanism may be related to increasing EphB2, reducing PARP 1 to curb excessive tau protein phosphorylation, protect the neuron dendritic spines and protect neurons function.