本文選題：阿爾茨海默病　切入點：固本健腦法　出處：《湖北中醫(yī)藥大學》2015年博士論文　論文類型：學位論文

【摘要】：目的隨著社會人口的快速老齡化,阿爾茨海默病(Alzheimer's disease, AD)因其高患病率及對社會、家庭的影響日益受到廣泛重視。因此,全世界均意識到加強研究AD的病因與發(fā)病機制,以及開發(fā)有效的防治措施和藥物的重要意義。本研究深入探討固本健腦法治療AD的作用機理,闡釋固本健腦治則治法理論的科學內涵,為尋求中醫(yī)改善學習記憶、治療AD的方藥提供依據(jù),豐富中醫(yī)固本理論及中醫(yī)防治AD理論的內容。方法1.理論研究：采用文獻研究方法,系統(tǒng)回顧中醫(yī)學從先后天之本認識老年性癡呆的理論,結合團隊相關研究成果入手,系統(tǒng)闡述固本健腦法治則治法理論。2.實驗研究：圍繞tau蛋白過度磷酸化損傷是AD發(fā)病的核心機制,以抑制tau蛋白過度磷酸化、保護樹突棘的信號傳導功能為靶點,選用安理申作為西藥對照,運用固本健腦法對AD大鼠模型進行干預,觀察其對AD模型的作用并探討其作用機制。取12月齡SPF級Wistar大鼠70只,雌雄各半。適應性喂養(yǎng)1w后,隨機分為7組,分別為正常組、空白對照組、模型組、固本健腦液高劑量組(簡稱固高組)、固中組、固低組和安理申組,每組10只。正常組不予處理。模型組和各治療組給予雙側海馬腦立體定位注射5μg/μl濃度的A β1-42：復制AD模型,空白對照組注射等位點等劑量生理鹽水。3d后,各治療組分別按照10ml/kg·d給予相應藥物濃度的固本健腦液和安理申混懸液灌胃,模型組與空白對照組則按照相同劑量給予生理鹽水灌胃,連續(xù)4w。灌胃結束后,對各組大鼠分別進行水迷宮定位航行實驗和空間探索實驗,觀察固本健腦法對AD大鼠模型行為學的影響。高爾基染色法觀察各組大鼠海馬神經(jīng)元樹突棘形態(tài)變化及密度改變。RT-PCR法檢測各組大鼠海馬EphB2、PARP-1水平變化。Western Blot法檢測各組大鼠海馬總tau、P-tau(Ser199, Ser396,Thr231位點)、EphB2、PARP-1表達變化。觀察固本健腦法對AD大鼠模型學習記憶能力、樹突棘形態(tài)及tau蛋白過度磷酸化的作用,以探討其防治AD的作用機制。結果1.固本健腦法對AD大鼠模型行為學的影響定位航行實驗中,模型組大鼠找到水下隱匿平臺潛伏期明顯長于正常組(P0.01),各治療組潛伏期明顯短于模型組(P0.01),且固高組、固中組潛伏期短于安理申組(P0.01)；目標象限游泳時間百分比比較,模型組明顯低于正常組(P0.01),各治療組明顯高于模型組(P0.01),且固中組高于安理申組(P0.05)�？臻g探索實驗中,模型組大鼠找到原平臺潛伏期明顯長于正常組(P0.01),各治療組潛伏期明顯短于模型組(PO.01),且固高組、固中組潛伏期短于安理申組(P0.05)；模型組大鼠穿越原平臺位置次數(shù)明顯少于正常組(P0.01),各治療組潛伏期明顯多于模型組(P0.01),且固高組、固中組多于安理申組(P0.01)；目標象限游泳時間百分比比較,模型組明顯低于正常組(P0.01),各治療組明顯高于模型組(P0.01),且固高組、固中組高于安理申組(P0.05)。表明本研究的AD模型是成功的,固本健腦法能明顯改善AD模型大鼠的學習記憶功能。2.固本健腦法對AD大鼠模型海馬神經(jīng)元樹突棘的影響比較各組大鼠的神經(jīng)元形態(tài)學發(fā)現(xiàn),模型組大鼠海馬區(qū)神經(jīng)元樹突棘密度明顯低于正常組(P0.01),各治療組樹突棘密度明顯高于模型組(P0.01),且固高組、固中組均高于安理申組(P0.01)。表明固本健腦法能明顯改善AD大鼠海馬神經(jīng)元樹突棘的形態(tài)。3.固本健腦法對AD大鼠模型海馬區(qū)P-tau的影響Western blot法檢測各組大鼠海馬tau、P-tau并比較P-tau水平,結果模型組大鼠較正常組明顯升高(P0.01),各治療組明顯低于模型組(P0.01),且固高組、固中組均低于安理申組(P0.01)。4.固本健腦法對AD大鼠模型海馬EphB2表達的影響Western blot與RT-PCR法檢測均顯示：模型組大鼠海馬EphB2表達較正常組明顯降低(P0.01),各治療組大鼠海馬EphB2表達明顯高于模型組(P0.01),且固高組、固中組大鼠海馬EphB2表達均高于安理申組(P0.05)。5.固本健腦法對AD大鼠模型海馬PARP-1表達的影響Western blot與RT-PCR法結果均顯示：模型組大鼠海馬PARP-1表達較正常組明顯升高(P0.01),各治療組大鼠海馬PARP-1表達明顯低于模型組(P0.05),且固中組高于安理申組(P0.05)。結論1.AD的發(fā)病與脾腎關系密切,先后天之本脾腎虧虛、腦髓失養(yǎng),兼有痰瘀是其基本病機,固本健腦法是治療該病的有效治法。2.固本健腦法能明顯改善AD模型大鼠的學習記憶能力；提高海馬神經(jīng)元樹突棘密度；升高海馬EphB2表達,降低PARP-1水平,抑制tau蛋白過度磷酸化。3.固本健腦法能有效防治AD,改善臨床癥狀,其機制可能與升高海馬EphB2、降低PARP-1從而抑制tau蛋白過度磷酸化,保護神經(jīng)元樹突棘、保護神經(jīng)元功能有關。
[Abstract]:Objective with the rapid population aging society, Alzheimer's disease (Alzheimer's, disease, AD) because of its high prevalence and the society, the influence of the family has received more and more attention. Therefore, all over the world are aware of strengthening the etiology and pathogenesis of AD has important significance, effective prevention measures and drugs and the development. Study the physic treatment mechanism of AD, explains the physic treatment scientific connotation of treatment theory, to improve the learning and memory of TCM, provide the basis for prescription in the treatment of AD, the rich Chinese medicine theory and the theory of TCM Prevention and treatment of AD. 1. research methods: literature research method, system from the review of the theory of traditional Chinese medicine has a knowledge of dementia, with team related researches, systematically expounded the physic treatment theory.2. experimental study on the rule of law: focus on tau protein Hyperphosphorylation of damage is the core mechanism of AD, to inhibit tau phosphorylation and protect the signaling function of dendritic spines as the target, selection of Aricept as western medicine control, using the physic method intervention on AD rat model, observe the effect of the AD model and explore its mechanism. 70 rats, 12 month old SPF Wistar female. Feeding after 1W, were randomly divided into 7 groups, including normal group, blank control group, model group, the physic liquid in high dose group (referred to as Gugao group), solid solid group, low dose group and donepezil group, n = 10 only. The normal group were not treated. The model group and the treatment group was treated with bilateral hippocampal brain stereotaxic injection of 5 mu g/ Mu l concentration of A beta 1-42: duplicate AD model, blank control group was injected with saline sites after.3d in different treatment groups were given corresponding drugs according to the 10ml/kg D concentration of the physic Liquid and Aricept suspension gavage, model group and blank control group with the same dose of saline, continuous intragastric administration of 4w. after the end of each rat were water maze navigation experiment and space exploration experiment, the physic method in observation of AD rat model for learning effects were observed in rat hippocampal neurons and morphological changes of dendritic spine density changes detected by.RT-PCR EphB2 in the hippocampus of rats of Golgi staining, PARP-1 level changes of.Western Blot tau were detected in the hippocampus of rats, P-tau (Ser199, Ser396, Thr231, EphB2 loci), expression of PARP-1. The physic method was used to observe the learning and memory the ability of AD rat model, effect of dendritic spine morphology and hyperphosphorylation of tau protein and to investigate its mechanism of prevention and treatment of AD. The effect of 1. methods to model the physic behavior of AD rats. In the experiment, the model group The rat found the latency of underwater is significantly longer than the normal group (P0.01), the treatment group was significantly shorter than that in the model group (P0.01), and Gugao group, solid group was shorter than Aricept group (P0.01); compare the percentage of swimming time in the target quadrant, the model group was significantly lower than normal group (P0.01), the treatment group was significantly higher than the model group (P0.01), and the solid is higher than the Aricept group (P0.05). Space exploration experiment, the rats in the model group to find the original platform was significantly longer than the normal group (P0.01), the treatment group was significantly shorter than that in the model group (PO.01), and high solid solid group. The incubation period is shorter than the Aricept group (P0.05); model group rats crossing the original platform number less than the normal group (P0.01), the treatment group was significantly more than the model group (P0.01 group), and high solid, solid group than Aricept group (P0.01); compare the percentage of swimming time in the target quadrant. Die Type group was significantly lower than normal group (P0.01), the treatment group was significantly higher than that in the model group (P0.01), and solid solid is higher than the high group, the Aricept group (P0.05). This study shows that the AD model is successful, the physic method can obviously improve the physic method of learning and memory function of.2. AD model the effects of rat hippocampal neurons of AD rat model of dendritic spine morphology of neurons were compared in rats found that neurons in hippocampus of model group rats spine density were significantly lower than the normal group (P0.01), the treatment group of spine density was significantly higher than the model group (P0.01), and solid solid group were higher than the high group. The Aricept group (P0.01) showed that the effect of the physic method. The morphology of.3. the physic method can obviously improve the hippocampal neuronal dendritic spines in AD rat model of P-tau AD in hippocampus of rats with Western blot method to detect the rat hippocampal tau, P-tau and P-tau level, results in the model group Rats increased significantly compared with normal group (P0.01), the treatment group was significantly lower than that of model group (P0.01), and solid solid group were lower than the high group, Aricept group (P0.01.4.) to detect the physic method on the expression of EphB2 in hippocampus of AD rat model and the effect of Western blot RT-PCR method showed that: the expression model of EphB2 in the hippocampus of rats significantly decreased compared with normal group (P0.01), EphB2 in hippocampus of rats in each treatment group was significantly higher than that in the model group (P0.01), and high solid solid group, expression of EphB2 in hippocampus of rats were higher than that of the Aricept group (P0.05) and Western blot RT-PCR method.5. the physic method on the expression of hippocampus PARP-1 AD rat model showed that the expression of PARP-1 in hippocampus of model rats increased significantly compared with normal group (P0.01), PARP-1 in hippocampus of rats in each treatment group was significantly lower than the model group (P0.05), and the solid is higher than the Aricept group (P0.05). Conclusion the incidence of 1.AD close relationship with spleen and kidney has A deficiency of spleen and kidney medulla dystrophy, both phlegm and blood stasis is the basic pathogenesis, the physic method is the physic method in treating the disease effective treatment.2. can significantly improve the learning and memory ability of AD rats; improve the hippocampal neuron dendritic spines; increased the expression of EphB2 in hippocampus, decreased the level of PARP-1. Inhibition of tau hyperphosphorylation of the physic method.3. can effectively control AD, improve the clinical symptoms, the mechanism may be related to increased hippocampal EphB2, decreased PARP-1 and inhibited tau phosphorylation and protection of dendritic spines, protect neuron function.

【學位授予單位】：湖北中醫(yī)藥大學
【學位級別】：博士
【學位授予年份】：2015
【分類號】：R285.5;R-332

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