【作者】 胡久略；

【导师】 贺又舜；

【作者基本信息】 湖南中医药大学 ， 方剂学， 2011， 博士

【摘要】 目的：通过复制反复性缺血再灌注VD模型,观测补肾醒脑方(BSXNP)对血管性痴呆(VD)模型大鼠学习记忆、脑组织病理形态学,脑组织炎性细胞因子TNF-α、IL-1β、IL-10、TGF-β,黏附分子ICAM-1、VCAM-1. P-selectin、E-selectin,胆碱能抗炎通路中CHAT、AchE与炎症因子的关系及NF-K B变化的影响,研究BSXNP对VD模型大鼠炎性反应的调控作用,进一步探讨BSXNP防治VD的作用机理。方法：第一部分：BSXNP对VD模型大鼠行为学及脑组织病理形态学的影响实验以反复缺血再灌注致VD模型大鼠为研究对象,以喜得镇组为对照组,将雄性Wistar大鼠随机分为模型组(MX)、假手术组(JS)、喜得镇组(XD)补肾醒脑方组(BX),用跳台法、水迷宫法分别于术后第7d、15d、30d观察各组大鼠学习记忆能力；于行为学实验结束后第二天,断头取大鼠脑组织,经HE染色观察各组大鼠脑组织病理形态学变化。第二部分：BSXNP对VD模型大鼠炎性反应的阻抑作用实验以反复缺血再灌注致VD模型大鼠为研究对象,以喜得镇组为对照组,将雄性Wistar大鼠随机分为MX、JS、XD、BX,于术后24小时取脑组织,运用ELISA法检测炎性细胞因子TNF-α、IL-1β、IL-10、TGF-β,黏附分子ICAM-1、VCAM-1、P-selectin、E-selectin含量,运用比色法检测CHAT、AchE活力,运用蛋白印迹法检测NF-κB表达。结果：第一部分：BSXNP对VD模型大鼠行为学及脑组织病理形态学的影响1.BSXNP能明显改善VD模型大鼠的学习记忆能力。BSXNP能明显缩短VD模型大鼠不同时间段跳台实验中触电反应期,延长触电潜伏期(P<0.01)；同时均可明显减少VD模型大鼠不同时间段学习和记忆成绩中的错误反应次数(P<0.01)；还可缩短VD模型大鼠在Morris水迷宫中寻找平台潜伏期,增加跨越平台次数(P<0.01或P<0.05)。且BSXNP组明显优于喜德镇组,差异具有统计学意义(P<0.05)。2.BSXNP能明显改善或减轻VD模型大鼠脑组织神经元的损伤,对脑神经细胞具有保护作用。第二部分：BSXNP对VD模型大鼠炎性反应的阻抑作用1.BSXNP能显著降低VD模型大鼠脑组织促炎因子TNF-α、IL-1β的含量(P<0.05),明显降低抗炎因子IL-10、TGF-β的含量(P<0.05)。2.BSXNP能不同程度的降低VD模型大鼠脑组织粘附分子ICAM-1、VCAM-1、E-selectin、P-select in含量(P<0.05或P<0.01),减轻缺血再灌注损伤。3.BSXNP能够提高VD模型大鼠脑组织ChAT活性(P<0.05),降低脑组织AchE活性(P<0.01),提示其可能亦作用于“迷走神经—胆碱能抗炎通路”,通过缓解VD模型大鼠迷走神经受到的抑制作用,使Ach的合成增多,减轻炎性反应,从而减少脑组织功能受损。4.BSXNP能抑制脑缺血再灌注后核转录因子-κB的的转录活性,下调NF-κB的表达,阻断炎症反应信号转导通路,减轻炎性损伤。结论：1.BSXNP能明显改善VD模型大鼠的学习记忆能力,减轻脑组织神经元的损伤,对脑神经细胞具有保护作用。2.本研究表明,在反复脑缺血再灌注后,炎症因子表达迅速上升,内皮细胞粘附分子增加,胆碱能抗炎通路未激活,核转录因子活化,从而引起脑组织损伤。给予BSXNP能阻断炎性反应信号转导通路,进而减轻炎症损伤。此表明脑缺血再灌注后出现的白细胞与内皮细胞粘附,炎性细胞聚集、浸润,炎症反应加重,微血管拥堵等是其组织病理基础之一；通过调控脑缺血再灌注后早期炎性反应是BSXNP治疗VD的重要作用机理之一,其可能机制是：BSXNP通过阻断NF-κB炎性反应信号转导通路和激活胆碱能抗炎通路,从而共同调节了炎性因子和黏附因子的表达,通过多通路、多靶点、多环节作用于炎症通路而实现调控炎性反应作用的。更多还原

【Abstract】 Objective:To observe the BSXNP’s influence on the relationship between VD model rat learning and memory,a brain biopsy morphology, and brain inflammatory cells factor TNF-α, IL-1β, IL-10, TGF-β, adhesion molecule ICAM-1, VCAM-1, P-selectin,E-selectin, and the influence on the change of the NF-κB; to study the inflammatory response in rats;to have a further discussing of the regulation function mechanism of action BSXNP prevention VD through the copy of repeatability ischemia-reperfusion.Methods:The first part:the BSXNP’s influence on the VD model of rats brain biopsy ethology and morphological.The experiment regard the repeated ischemia-reperfusion cause VD rat models as research object, regard the the joy of as control group, the male Wistar rat randomly is divided into model group (MX), jiaShouShu group (JS), XiDeZhen group (XD), Bu Shen Xing Nao Prescription group (BX), using the diving platform method and water maze method to observe the learning and memory capacity of rats in each group after the experiment respectively in the time of postoperative 7d, 15d,30d. On the second day of the finish of the experiment behaviouristics, bufo by rats organization, take each rats HE dyed observation histopathological morphological changes. Part2:the resistance inhibitory effect of the BSXNP to the inflammatory response of the rat models.The experiment regard the repeated ischemia-reperfusion cause VD rat models as research object, the joy of as control group, divided the male Wistar rat randomly into MX, JS, XD, BX,24hours after the operation, take out the brain by using ELISA take brain test inflammatory cells factor TNF-α, IL-1β, IL-10, TGF-β, adhesion molecule ICAM-1, VCAM-1, P-selectin E-selectin content, using colorimetric detection CHAT, AchE vigor, use protein imprinting method to detect NF-κB expression.Results:The first part:the influence of BSXNP on the VD model of rats brain biopsy ethology and morphological1. BSXNP can improve obviously the VD model rats learning and memory capacity obviously. BSXNP can shrink the response time of VD rat models in different period of the platform experiment obviously, extended the time of electric shock reaction (P<0.01). In the same time, it can also reduce times of the fault reaction in the learning and remembering grade of the VD rat model in the different period (P<0.01). It can also shorten VD model rat’time of searching platform in the Morris water maze and increase the times of acrossing platforms(P<0.01 or P<0.05). And BSXNP group’s obviously superior to the group, starwood a statistically significant difference (P<0.05). 2. BSXNP can significantly improve or mitigate the injury of the VD model rats’s neurons organizational and BSXNP has a protective effect of the brain nerve cells.Part 2:BSXNP has the resistance inhibitory effect on the inflammatory response of the VD rat models.1. BSXNP can significantly reduce VD model rats organization promote inflammation factors TNF alpha, IL-1βcontent (P< 0.05), decreased the content of anti-inflammatory factor IL-10, TGF-βobviously (P<0.05).2. BSXNP can reduce VD model rats organization ICAM-1, adhesion molecules VCAM-1, E-selectin, P-selectin content in different degree (P<0.01 or P<0.05), and lighten the injury of the chemia-reperfusion.3. BSXNP can improve the activity of ChAT of VD model rats (P<0.05), reducing the activity of AchE (P<0.01), suggesting that its may also play a role in the vagus nerve-choline can anti-inflammatory pathways, VD model rats by dampening the vagus nerve, inhibiting effect by increasing, reduce the synthesis of Ach, inflammatory reaction and thereby reducing brain dysfunction.4. BSXNP can restrain nuclear transcription factors-κB transcription of activity after the brain ischemia reperfusion, reduce NF-κB expression, blocking inflammation, relieve signal transduction pathways inflammatory lesions. Conclusion:1. BSXNP can obviously improve the VD model rats the ability of learning and memory in, reduce the damage of brain neurons, protectie effect of nerve cells.2. This study shows that, after the repeated cerebral ischemia and reperfusion, the inflammatory factor expression rising rapidly, endothelial cell adhesion molecules, choline increased, the anti-inflammatory pathways were not be activated, nuclear transcription factors become active and lead to damage of brain. BSXNP be given to pathways which can block the inflammatory reaction signal, and thus can alleviate inflammation damage. This shows that cerebral ischemia reperfusion appeared after the leucocyte and endothelial cell adhesion, inflammatory cell aggregates, infiltration, inflammation is aggravating, such is its organization capillaries congestion through regulation, one of the pathology base after cerebral ischemia and reperfusion BSXNP treatment early inflammatory reaction is one of the important role the mechanism.Its possible mechanism is: BSXNP by blocking NF-κB inflammatory reaction signalling pathways, and activate the choline can anti-inflammatory pathways, thus common adjust inflammatory factor and adhesion factor expression through multiple access, more and more targets, in the role of inflammatory pathways and link to realize regulation of the inflammatory response function.更多还原