本文關(guān)鍵詞：健腦1方治療血管性癡呆療效觀察及對NMDAR-CaMKⅡ通路的影響　出處：《廣州中醫(yī)藥大學(xué)》2015年博士論文　論文類型：學(xué)位論文

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【摘要】：目的：血管性癡呆(Vascular Dementia, VD)是指缺血性、出血性腦血管疾病引起的腦損害導(dǎo)致的癡呆,它在各種老年期癡呆的病因中僅次于阿爾茨海默病(Alzheimer's disease, AD)和路易體癡呆(Dementia with Lewy body, DLB),約占老年期癡呆的20%左右。VD的患病率隨年齡增長呈指數(shù)上升,嚴(yán)重影響到患者的生存質(zhì)量,并給其家庭甚至社會帶來了沉重負(fù)擔(dān)。鑒于VD的可防治性,應(yīng)及早干預(yù)治療,預(yù)后良好。由于其病因及發(fā)病機制目前尚不清楚,且西藥不良反應(yīng)較多,運用中醫(yī)藥治療VD越來越受到人們的關(guān)注,近年來,國內(nèi)學(xué)者對中醫(yī)藥治療本病進行了深入研究,發(fā)現(xiàn)中醫(yī)藥治療VD的優(yōu)勢與特色在于可顯著改善臨床癥狀、減少不良反應(yīng)。國醫(yī)大師鄧鐵濤教授根據(jù)自己多年的臨床經(jīng)驗,提出以“益氣除痰活血法”治療血管性癡呆的學(xué)術(shù)思想,鄧?yán)险J(rèn)為本病主要病機是氣虛為本,痰瘀為標(biāo),由痰致瘀,以痰為主。治療上牢抓中醫(yī)辨證論治的“整體觀”,認(rèn)為本病雖病位在腦,但發(fā)病之根本在于脾,故強調(diào)本病治療關(guān)鍵在于益氣健脾,并主張治療痰瘀同病時重在化痰,輔以祛瘀,自擬健腦1方在臨床實踐中療效顯著,值得深入研究及臨床推廣。為了更好地總結(jié)鄧鐵濤教授臨床經(jīng)驗,探究其臨證規(guī)律及學(xué)術(shù)特色,挖掘其科學(xué)內(nèi)涵,冀以弘揚鄧?yán)蠈W(xué)術(shù)思想,傳承仁術(shù)。本課題是在中醫(yī)理論的指導(dǎo)下,結(jié)合鄧鐵濤教授多年的臨床經(jīng)驗,客觀評價健腦1方治療VD的臨床療效及其安全性,為本方良好的臨床療效提供客觀數(shù)據(jù)支持,從而更好地繼承名老中醫(yī)學(xué)術(shù)經(jīng)驗,豐富中醫(yī)學(xué)理論,繼承和發(fā)揚中醫(yī)藥事業(yè)。并通過對大量國內(nèi)外文獻研究,提出“健腦1方通過干預(yù)NMDAR-CaMKⅡ信號傳導(dǎo)通路調(diào)控一系列下游蛋白是其抗VD的可能作用機制”的假設(shè)。通過對VD大鼠海馬NR2B和CaMKⅡ的基因表達深入研究有望闡明健腦1方影響VD大鼠的學(xué)習(xí)記憶能力主要的分子生物學(xué)作用機制,并為中醫(yī)藥治療VD的研究提供新的靶點和思路。方法：臨床研究：本課題采用平行、隨機、對照實驗方案,選擇2014年1月至2014年12月就診于廣州中醫(yī)藥大學(xué)第一附屬醫(yī)院心內(nèi)科及廣東省第二中醫(yī)院腦病科門診及住院符合血管性癡呆(氣虛痰瘀證)診斷標(biāo)準(zhǔn)患者60例,采用隨機數(shù)字表法按照1：1的比例將60例患者隨機均分成治療組、對照組各30例,同時給予兩組患者降壓、降脂等一般治療措施,其中治療組給予健腦1方治療,對照組予尼莫地平片治療,治療期間停用其它與治療本病有關(guān)的內(nèi)服藥物,療程共6周。治療前與治療6周后記錄每例入組患者的MMSE評分、ADL評分、NPI評分、中醫(yī)證候積分的變化,并記錄治療前后血常規(guī)、肝腎功能等安全性指標(biāo)與治療相關(guān)的不良反應(yīng)事件,以客觀準(zhǔn)確評價健腦1方治療血管性癡呆的臨床療效及安全性。實驗研究：本研究采用雙側(cè)頸總動脈永久性結(jié)扎建立VD動物模型,將60只SD雄性大鼠隨機分為假手術(shù)組、模型組、高劑量組、低劑量組及尼莫地平組,低、高劑量組、尼莫地平組大鼠分別灌服低劑量健腦1方、高劑量健腦1方和尼莫地平混懸液；模型組、假手術(shù)組均給予生理鹽水灌胃,給藥30天,用Morris水迷宮檢測各組大鼠的學(xué)習(xí)記憶能力,HE染色觀察大鼠海馬組織學(xué)改變,用RT-PCR法檢測大鼠海馬NR2BmRNA及CaMKⅡmRNA表達水平,從而探討鄧鐵濤教授健腦1方對血管性癡呆大鼠的學(xué)習(xí)記憶能力的影響,并從分子生物學(xué)角度探討其可能的作用機制。成果：臨床研究：試驗結(jié)果顯示：兩組患者在性別、年齡、病程、病情及文化程度方面等因素構(gòu)成比的差異性無統(tǒng)計學(xué)意義(P0.05),提示各組患者在性別、年齡、病程、病情及文化程度的構(gòu)成方面基本均衡,具有可比性。兩組患者治療前MMSE、ADL、NPI及中醫(yī)證候積分均未見有統(tǒng)計學(xué)差異(P0.05),具有可比性；最終研究結(jié)果可見：經(jīng)過6周治療,兩組患者治療后MMSE評分均較治療前提高,具有顯著性差異(P0.01)；治療后兩組患者MMSE評分比較無顯著性差異(P0.05),兩組治療前后差值比較差異有統(tǒng)計學(xué)意義(P0.01)；治療組總有效率80.0%,對照組總有效率53.3%,提示兩組均能改善患者認(rèn)知能力,且治療組優(yōu)于對照組(P0.05)。兩組患者治療后ADL評分均較治療前降低,具有顯著性差異(P0.01)；治療后兩組患者ADL評分及兩組治療前后差值比較均無顯著性差異(P0.05)；治療組總有效率23.3%,對照組總有效率30.0%,提示兩組均能提高患者日常生活活動能力,且臨床療效相近(P0.05)。兩組患者治療后NPI評分與治療前比較均無顯著性差異(P0.05)；治療后兩組患者NPI評分及兩組治療前后差值比較均無顯著性差異(P0.05)；治療組總有效率6.67%,對照組總有效率6.67%,提示兩組均未能顯著改善患者精神行為癥狀(P0.05),且兩組均不能有效減少VD患者照料者的苦惱程度(P0.05)。治療組患者治療后中醫(yī)證候積分較治療前降低,具有顯著性差異(P0.01),對照組患者治療后中醫(yī)證候積分與治療前比較無顯著性差異(P0.05),治療后兩組患者中醫(yī)證候積分及兩組治療前后差值比較均有顯著性差異(P0.01)；治療組總有效率為56.67%,對照組總有效率為10.0%,提示治療組可顯著改善患者中醫(yī)證候,對照組患者中醫(yī)證候改善甚微(P0.01)。參加本次研究的所有患者均未出現(xiàn)血常規(guī)及肝腎功能檢查異常,兩組患者出現(xiàn)的不良反應(yīng)癥狀輕微,安全性好。實驗研究：Morris水迷宮測試結(jié)果：隨著訓(xùn)練天數(shù)的增加,各組大鼠逃避潛伏期逐漸縮短,說明各組大鼠在學(xué)習(xí)訓(xùn)練中逐漸學(xué)會尋找平臺,但學(xué)習(xí)記憶能力存在顯著差異,其中訓(xùn)練四天內(nèi)每一天模型組大鼠逃避潛伏期均較假手術(shù)組明顯延長(P0.05或P0.01),說明模型組大鼠空間學(xué)習(xí)記憶能力出現(xiàn)障礙,第三天,高劑量組大鼠逃避潛伏期較模型組明顯下降(P0.05)；第四天,高劑量組及尼莫地平組大鼠逃避潛伏期均較模型組顯著下降(P0.01和P0.05)。空間探索實驗結(jié)果顯示：120s內(nèi),高劑量組及假手術(shù)組大鼠穿越平臺的次數(shù)明顯高于模型組,差異具有統(tǒng)計學(xué)意義(P0.01)；各組大鼠停留在平臺所在象限的游泳時間與模型組大鼠比較差異尚未發(fā)現(xiàn)統(tǒng)計學(xué)意義(P0.05),由此可見,實驗結(jié)果說明高劑量組VD大鼠學(xué)習(xí)記憶能力顯著提高。VD大鼠海馬組織學(xué)改變：光鏡下觀察發(fā)現(xiàn)模型組大鼠海馬神經(jīng)細(xì)胞丟失；神經(jīng)細(xì)胞核碎裂、核膜不清、核仁消失,神經(jīng)細(xì)胞收縮,胞漿嗜酸性增強,膠質(zhì)細(xì)胞反應(yīng)性增生；病變集中在CA1、CA2及CA3區(qū)；尼莫地平組大鼠海馬病變集中在CA1區(qū),有神經(jīng)元細(xì)胞損傷；高、低劑量組大鼠海馬病變集中在CA1、CA3區(qū),可見神經(jīng)元細(xì)胞損傷,病變程度較模型組減輕,提示健腦1方及尼莫地平可減少VD大鼠海馬組織神經(jīng)元損傷范圍及程度。大鼠海馬NR2B mRNA及CaMKⅡmRNA表達：實驗結(jié)果表明模型組大鼠海馬NR2B mRNA的表達水平與假手術(shù)組相比明顯降低(P0.01)。而與模型組相比較,尼莫地平組及高劑量組大鼠海馬NR2B mRNA的表達水平顯著增高,差異具有統(tǒng)計學(xué)意義(P0.05),低劑量組有增高NR2B mRNA表達的趨勢。模型組大鼠海馬CaMKⅡmRNA的表達與假手術(shù)組相比稍降低(P0.05),高劑量及尼莫地平組都能略微提高mRNA的表達水平,但差異無均統(tǒng)計學(xué)意義(P0.05)結(jié)論：臨床研究結(jié)果提示健腦1方治療VD在改善患者認(rèn)知功能、提高患者日常生活活動能力方面具有良好療效,還能明顯改善患者的中醫(yī)臨床癥狀,但對VD患者精神行為癥狀未見明顯改善,且不良反應(yīng)少,安全性好。本課題研究印證了基于鄧?yán)咸岢龅摹耙鏆獬祷钛ā敝委熝苄园V呆理念的正確性和科學(xué)性,以及基于鄧?yán)吓R床經(jīng)驗而創(chuàng)制的健腦1方的有效性和安全性。實驗研究結(jié)果提示健腦1方可顯著提高VD大鼠的學(xué)習(xí)記憶能力,并推測其作用機制可能與減少VD大鼠海馬組織神經(jīng)元損傷范圍及程度,并可上調(diào)VD大鼠海馬NR2B mRNA的表達水平有關(guān)。本實驗發(fā)現(xiàn)VD大鼠海馬CaMKⅡmRNA并沒有得到同樣的改善,提示CaMKⅡ可能不是健腦1方的重要作用靶點。
[Abstract]:Objective: vascular dementia (Vascular Dementia, VD) refers to ischemic brain damage, dementia, hemorrhagic cerebrovascular disease caused by it, after Alzheimer's disease in the etiology of senile dementia in (Alzheimer's disease AD) and Louis (Dementia with Lewy body body dementia, DLB), accounting for about old age 20%.VD the prevalence of dementia increased exponentially with the increase of age, seriously affect the patient's quality of life, and to their families and society a heavy burden. In view of prevention and treatment of VD, should be early intervention, the prognosis is good. Its etiology and pathogenesis is unclear, and many adverse reactions of Western medicine the use of Chinese medicine treatment of VD, more and more attention in recent years, domestic scholars on Chinese medicine treatment of this disease is studied, found the advantages and characteristics of traditional Chinese medicine in the treatment of VD is significantly improved by The clinical symptoms, reduce adverse reaction. The Chinese medicine master professor Deng Tietao according to their clinical experience for many years, put forward the idea of "Invigorating Qi and removing phlegm and activating blood" in the treatment of vascular dementia academic thought, Denglao thinks that its pathogenesis is the deficiency of Qi, phlegm stasis, the phlegm and blood stasis, phlegm in the treatment. Firmly grasp the dialectic of "wholeness", believe that although this disease disease in the brain, but the pathogenesis lies in the spleen, it emphasizes the treatment of this disease lies in Qi and spleen, and advocate the treatment of phlegm and blood stasis phlegm and removing blood stasis, when focusing on, with brain 1 party in clinical practice has obvious curative effect, worthy of further study and clinical application. In order to summarize the clinical experience of Professor Deng Tietao, to explore the clinical characteristics and academic characteristics, to explore its scientific connotation, in order to promote the academic thoughts of Deng, Renzhu inheritance. This paper is under the guidance of Chinese medicine theory, combined with Deng Tietao taught "Clinical experience for many years, the objective evaluation of 1 Jiannao Decoction on VD clinical curative effect and safety, to provide objective data support for the good clinical curative effect, so as to better inherit the old Chinese academic experience, enrich the theory of traditional Chinese medicine, inheritance and development of traditional Chinese medicine. Based on a large number of domestic and foreign literature research, put forward" 1 party intervention regulation by brain NMDAR-CaMK II signal transduction pathway downstream protein is its anti VD mechanism "hypothesis. Further research is expected to elucidate the molecular mechanism of brain 1 affect the ability of learning and memory in VD rats by mainly on VD rat hippocampal NR2B and CaMK II gene expression, and provide a new target and idea for the research of Chinese medicine for the treatment of VD. Methods: clinical study: this subject adopts a parallel, randomized, controlled experiment scheme, from January 2014 to December 2014 in Guangzhou Encephalopathy outpatient department of Cardiology the First Affiliated Hospital and the second hospital of Guangdong Province, University of traditional Chinese medicine and hospital with vascular dementia (Qi deficiency and phlegm and blood stasis syndrome) 60 cases of patients, randomly according to the ratio of 1:1 60 patients were randomly divided into treatment group and control group with 30 cases in each group, and given two patients with hypertension, cholesterol and other general treatment measures, the treatment group was treated with 1 brain decoction, control group received Nimodipine Tablets treatment, during treatment discontinuation and other treatment of the disease related to oral drugs, the treatment lasted for 6 weeks. Before treatment and after 6 weeks of treatment were recorded in groups of patients with MMSE score, ADL score, NPI score and the changes of TCM syndrome integral, and record the blood before and after treatment, liver and renal function indicators of adverse events associated with treatment, to objectively evaluate the clinical curative effect of 1 treatment of brain vascular dementia and ANN Full. Experimental research: animal model of VD was established by permanent bilateral common carotid artery ligation by this study, 60 male SD rats were randomly divided into sham operation group, model group, high dose group, low dose group and nimodipine group, low and high dose group, nimodipine group rats were fed low dose brain 1 Party, the 1 party and the high dose of Jiannao nimodipine suspension; model group, sham operation group were given normal saline, administered for 30 days, with the ability of learning and memory in Morris water maze test in rats, observe changes in hippocampus of rats with HE staining, RT-PCR assay in hippocampal NR2BmRNA and CaMK II mRNA expression level, so as to explore the influence of the ability of learning and memory of Professor Deng Tietao, the 1 Party of brain vascular dementia rats, and to explore its possible mechanism from the perspective of molecular biology. Results: the clinical study: the experimental results showed that the patients in the two groups of No, age, duration of illness, and the degree of cultural factors such as the ratio difference was not statistically significant (P0.05), suggesting that patients in gender, age, course of disease, the basic balance of composition condition and culture degree, comparable. Before MMSE, ADL two groups, NPI and TCM syndrome there were no statistically significant differences between the integral (P0.05), comparable; the final research results showed: after 6 weeks of treatment, improve the MMSE score of the two groups were lower than before treatment, with significant difference (P0.01); after treatment, MMSE scores between two groups had no significant difference (P0.05), there was statistical the significance of the two groups before and after treatment (P0.01); the difference between the treatment group total effectiveness 80%, control group total effective rate was 53.3%, suggesting that the two groups can improve the cognitive ability of the patients, and the treatment group than the control group (P0.05). The two groups of patients ADL score after treatment than before treatment Decreased, with significant difference (P0.01); after treatment, two groups of patients before and after treatment in the two groups of ADL score and the difference had no significant difference (P0.05); the total effective rate of treatment group 23.3%, control group total effective rate was 30%, suggesting that the two groups could improve the activities of daily living of patients, and the clinical curative effect is similar to (P0.05). NPI score of the two groups compared with before treatment showed no significant difference (P0.05); after treatment, two groups of patients before and after treatment in the two groups of NPI score and the difference had no significant difference (P0.05); the total effective rate of treatment group 6.67%, control group total effective rate was 6.67%, suggesting that the two groups no significant improvement in patients with behavioral and psychological symptoms (P0.05), and the two groups can not effectively reduce the distress of patients with VD caregivers (P0.05). The patients in the treatment group after treatment of TCM syndrome score lower than before treatment, with significant difference (P0.01), patients in the control group after treatment TCM syndrome score before treatment showed no significant difference (P0.05), two groups of patients before and after the TCM syndromes and treatment in the two groups the difference had significant difference after treatment (P0.01); the total efficiency of treatment group was 56.67%, control group total effective rate was 10%, indicating that the treatment group of patients with TCM syndrome significantly, the control group of TCM syndromes in patients with poor effect (P0.01). All patients in this study were not found in blood and liver function abnormalities and adverse reactions of two groups of patients with mild symptoms and good safety. Experimental research: Morris water maze test results: with the increase in the number of training days, rats the escape latency shortened gradually, that the rats in learning and training gradually learn to find the platform, but there are significant differences between the ability of learning and memory, including training four days every day the rats in the model group compared to the sham operation latency time Group significantly increased (P0.05 or P0.01), the model group of spatial learning and memory abilities of rats appeared disorder, third days, the escape latency of rats in high dose group decreased significantly compared with model group (P0.05); the fourth day, the high dose group and nimodipine group rats escape latency were significantly decreased compared with the model group (P0.01 and P0.05). Space exploration results showed: in 120s, the number of high dose group and sham operation group rats through the platform was significantly higher than the model group, the difference was statistically significant (P0.01); the rats staying in the platform quadrant and the rats in the model group the difference has not been found statistically significant (P0.05), thus the experimental results show that the high dose group of VD rats learning and memory ability to significantly improve the pathological changes of hippocampus in.VD rats: Lost nerve cells in the hippocampus of model group rats were observed under light microscope; nerve nuclei fragmentation, nuclear membrane Not clear, nucleolus disappeared, nerve cell shrinkage, cytoplasmic eosinophilic enhancement, astrocytes proliferation; lesion concentration in CA1, CA2 and CA3 region of hippocampus lesions; nimodipine group rats were concentrated in the CA1 area, a neuronal cell injury; high, low dose group rat hippocampal lesion concentration in CA1, CA3 District visible, neuronal cell injury, lesions alleviated as compared with model group, suggesting that the brain of 1 party and nimodipine can reduce the range and extent of neuronal damage in the hippocampus of VD rats. The expression of NR2B in hippocampus mRNA and CaMK II mRNA: the experimental results showed that the expression level of NR2B in hippocampus of mRNA model rats were significantly decreased compared with the sham operation group (P0.01). Compared with the model group, the expression level of nimodipine group and high dose group rat hippocampal NR2B mRNA increased significantly, the difference was statistically significant (P0.05), low dose group increased NR2B mRNA expression in the model group. The expression of CaMK in hippocampus of mRNA was slightly decreased compared with the sham operation group (P0.05), high dose group and nimodipine can slightly improve the expression level of mRNA, there were statistically significant but the difference (P0.05) conclusion: the clinical results suggest that brain 1 treatment of VD patients in improving the cognitive function, has good curative effect to improve the ability of daily life of patients, can significantly improve patients' clinical symptoms, but significantly improved for patients with behavioral and psychological symptoms were not VD, and less adverse reaction, good safety. This research confirms based on Deng's "Invigorating Qi and removing phlegm and activating blood" in the treatment of vascular dementia of the correct and scientific concept based on clinical experience, and Deng created brain 1 party is effective and safety. The experimental results suggest that the brain 1 party can significantly improve the learning and memory ability of VD rats, and its mechanism can be speculated It can be related to reducing the extent and extent of neuronal damage and increasing the expression level of NR2B mRNA in hippocampus of VD rats. It is found that the CaMK II mRNA in hippocampus of VD rats has not been improved similarly, suggesting that CaMK II may not be an important target for the 1 Party of brain strengthening.

【學(xué)位授予單位】：廣州中醫(yī)藥大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R277.7

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