Effect Of Compatibility Of Active Components In Acorus Gramineus On Learning And Memory And Relevant Mechanism In APPswe/PS1dE9Double Transgenic Mice

BackgroundAlzheimer’ s disease (AD) is a common type of dementia and neurodegeneration diseases. Age and dementia family history are two risk factors for AD pathogenesis. With the aging of the world’s population, there will be higher prevelance of AD, bringing heavy social and economic burden. So it becomes of great significance and prospects for further research and development of therapy for AD.Until now, etiology and pathogenesis of AD is unclear. Neural molecular pathology research suggests that AD is the abnormal expression and processing of the protein. The amyloid-protein (beta-amyloid, of A β) misfolding, aggregation and deposition in the senile plaques of AD pathological signs. Aβ toxic damage to neurons has been research focus of AD. Recent studies suggest that insulin signaling pathway and Aβ metabolism are closely related, insulin signaling pathway play an important role in protection of neurons in AD patients.Chinese medicine has rich theory and effective prescription in the treatment of dementia. Acorus gramineus is one commonly used Chinese Medicine in the treatment of dementia. Related research already verified the therapeutic effect in vivo and in vitro model of main active ingredient beta-asarone, eugenol of Acorus gramineus. The best ratio of beta-asarone, eugenol compatibility-4:1, has also been identified. On the basis of preliminary study, this experiment will further explore the mechanism of acorus gramineus active ingredient compatibility on Aβ metabolism and neuroprotective effects, using APPswe/PS1dE9double transgenic mouse model. ObjectivesThrough compareing the effect of acorus gramineus active components compa-tibility soup (β-asarone and eugenol in4:1) and β-asarone on learning memory, Aβ metabolism, insulin sinaling pathways and apoptotic signals of APPswe/PS1dE9mice, to explore mechanisem of acorus gramineus active components compatibility and β-asarone in curing AD, and to explore whether compatibility can have better effect and less toxicity. Then to provide a theoretical basis for the development of prevention and treatment of AD in traditional Chinese medicine.Methods1.To observe the changes of Aβ1-42area-. IGF-1R and apotosis in5months,7months and10months APPswe/PSldE9mice, using immunohistochemistry and Tunnel method.2. Male APPswe/PS1dE9double-transgenic mice were divided randomly into model group,β-asarone treated(84.8mg/kg per day) group, compatibility of low-dose group(42.4mg/kg per day; β-asarone:eugenol=4:1), compatibility of high-dose group(84.8mg/kg per day; β-asarone:eugenol=4:1) and donepezi treated(4mg/kg per day) group,19mice per group. In addition,19male with the same age and background APPswe/PS1dE9negatively transgenic mice were used as normal control group. Distilled water was administered to the normal control group and model group. All mice were oral administered by the drugs from5to10months of age. After5months of treatment, spatial learning and memory was measured by Morris Water Maze.3.To observe the change of APP、BACE1and Aβ42in cerebral cortex of APPswe/PS1dE9mice using Western-blot methods, to observe senile plaques in cerebral cortex using immunohistochemistry and Thioflavin-S staining method, then to evaluate the therapeutic effects of compatibility of active components in acorus gramineus on Aβ cascade reactions.4. To observe the change of IGF-1R/p-IGF-1R、AKT/p-AKT and Bcl-2in cerebral cortex of APPswe/PS1dE9mice using Western-blot methods, to observe the change of apoptosis using Tunnel methods and evaluate the therapeutic effects of compatibility of active components in acorus gramineus on insulin signal transduction and apoptosis. Rusult1. The immunohistochemical results suggest that there is a certain amount of senile plaques deposition in5months old APPswe/PS1dE9transgenic mice.7months,10months mice can be seen a large number of senile plaques,have significant difference compare to5months (P<0.05). Senile.plaques are mostly-concentrated in the parts of the cortex. Compared to5months and7months APPswe/PS1dE9transgenic mice, IGF-1R positive cell in10moths mice significantly increased (P<0.05). Tunnel apoptosis prompt apoptosis increased by months of age growth in mice.2. In Morris water maze test, escape lantacy of model group were significantly increasedand and the times of crossing the platform reduced significantly compared to normal group (P<0.05);Compared to model group, β-asarone treated group and compatibility of high-dose group show shorten escape lantacy(P<0.05);There is on difference between β-asarone treated group and compatibility of high-dose group.3. Western blot results showed that, Compared with normal group mice, the expression-of APP, BACE1and Aβ1-42were significantly increased in model group(P<0.05). Compared to model group, all groups that give drugs show the trend to reduce the expression of BACE-1and Aβ1-42protein, but the rusult was not statistically significant (p>0.05). Immunohistochemistry and sulfur staining results show that, senile plaques of the model group increased significantly compared with normal group, and treated group and model group were no significant differences.4. Western blot results showed, compared to normal group, IGF-IR protein of model group increased significantly(P<0.05).Compared to model group, donepezi treated group, β-asarone treated group, compatibility of low-dose group decrease IGF-1R significantly (P<0.05).The expression of p-IGF-1R、 AKT/p-AKT have no significant difference. Compared to normal group, the expression of Bcl-2of model group decreasede significantly (P<0.05), and Bcl-2increased significanty in β-asarone treated group, compatibility of low-dose group and compatibility of high-dose group compared to model group (P<0.05) The TUNNEL detection results suggest that the β-asarone groups, the compatibility group of low-dose and high dose compatibility group can be significantly reduced apoptosis compared with model group (P<0.05)Conelusion1. senile plaques increased significantly with age in APPswe/PS1dE9mice, accompanied part of the.insulin signal transduction pathway protein expression abnormal. The expression of apoptosis cells has been expressed since the5-month-old transgenic mice.2. β-asarone treated group(β-asarone:84.8mg/kg/d) and compatibility of high-dose group(β-asarone:67.84mg/kg/d, eugenol:16.96mg/kg/d) can effectively improve learning and memory of APPswe/PS1dE9mice. Compared to β-asarone treated group, compatibility group does not show the advantage of "reduce toxicity and increase efficiency"3. Compatibility of active components in acorus gramineus can not effectively influence the on Aβ cascade reactions in APPswe/PSldE9mice.4. Compatibility of active components in acorus gramineus can not improve the expression of upstream protein. molecules of insulin signal transduction pathway, but can up-regulating Bcl-2expression and reduced apoptosis.5.The mechanism about inhibition of neuronal apoptosis of compatibility of active components in acorus gramineus needs further study.