辣椒素通过激动TRPV1受体对帕金森病大鼠模型神经保护作用研究

发布时间：2018-06-04 12:20

本文选题：辣椒素 + TRPV1　；参考：《山东大学》2017年硕士论文

【摘要】：研究背景和目的黑质致密部多巴胺能神经元变性死亡所导致的体内多巴胺(DA)及其代谢产物含量减少是导致帕金森病的主要原因,其减少程度与多巴胺能神经元丧失程度成正相关。络氨酸羟化酶(TH)为DA合成限速酶,研究证明PD患者及PD动物模型TH含量明显降低,因此,通过干预提高TH含量及活性已成为减缓PD进展的重要治疗策略。瞬时受体电位香草醛亚型1(TRPV1)是一种非选择性阳离子通道,可被辣椒素选择性激活。传统上,TRPV1被认为涉及广泛的疾病领域,包括疼痛(炎症,内脏,癌症和神经病),炎症性肠病,间质性膀胱炎,尿失禁,气道疾病,胰腺炎和偏头痛,其涉及到的机制包括炎症,免疫,凋亡,氧化应激等诸多方面。最新研究表明TRPV1不仅在感觉神经元高度表达,而且高表达于其他功能脑区,并参与神经元损伤的细胞过程。此外,深入研究发现TRPV1可降低氧化应激以及脑梗死的相关标志物,提高运动和认知功能。提示其可能在神经退行性疾病的发生发展中发挥作用。本研究通过使用6-羟基多巴胺(6-OHDA)诱导的帕金森病大鼠模型来探讨TRPV1的神经保护作用,为其可能作为新的药物治疗靶点提供理论依据。研究方法随机选取108只雄性健康Wistar大鼠,体重控制为220g-250g,由山东大学动物实验中心提供。将其随机分为正常对照组(20只)、假手术组(右侧脑前内侧束立体定向注射生理盐水,20只),6-OHDA组(右侧脑前内侧束立体定向注射6-OHDA,40只),6-OHDA+辣椒素组(即干预组,注射6-OHDA后每天每只给予辣椒素1mg/kg腹腔注射,连续注射7d,辣椒素剂量为1mg/kg,40只)。对大鼠分别进行行为学检测(包括旋转试验与旷场试验)、蛋白质印迹检测黑质及纹状体TH及TRPV1表达水平、免疫组织化学检测黑质TH+神经元、测定大鼠脑内CAT、SOD、MDA 含量。所有实验数据均使用GraphPad Prism 5.0软件处理。计量资料以均数士标准差(x ±s)进行统计描述;采用t检验进行两组间均数比较,p0.05为差异具有统计学意义。研究结果1.行为学结果示与6-OHDA组相比,干预组大鼠诱发旋转圈数明显减少(n=10,p0.01),大鼠1分钟内行走距离明显增多,平均速度明显增快,中央格停留时间显著减少,且差别均具有统计学意义(n=10,p0.05)。2.免疫印迹结果示干预组大鼠黑质及纹状体TRPV1及TH蛋白表达水平较其余三组明显增高,差异均具有统计学意义(n=6,p0.01)。3.组化结果示与6-OHDA组相比,干预组黑质TH+神经元数目明显增多,此外干预组大鼠黑质致密部TH阳性神经元树突明显增多,轴突长度增加。4.与6-OHDA组相比,干预组右脑CAT、SOD水平均明显增高,MDA水平明显降低,差异均有统计学意义(t=2.977,p=0.0139;t=2.787,p=0.0192;t=2.563,p=0.0282),左脑6-OHDA组与干预组相比CAT、SOD、MDA水平差异无统计学意义(p0.05)。结论辣椒素可激活TRPV1蛋白,调节氧化应激水平,对PD大鼠多巴胺能神经元起神经保护作用,改善其运动功能,缓解PD病情。
[Abstract]:Background and objective the main cause of Parkinson's disease is the decrease of dopaminergic DAs and their metabolites in vivo due to the degeneration and death of dopaminergic neurons in the substantia nigra. The degree of decrease was positively correlated with the degree of loss of dopaminergic neurons. Th) is a rate-limiting enzyme for DA synthesis. It has been proved that the content of th in PD patients and PD animal models is significantly decreased. Therefore, increasing the content and activity of th through intervention has become an important therapeutic strategy to slow down the progress of PD. Transient receptor potential vanillin subtype 1 (TRPV1) is a nonselective cationic channel, which can be selectively activated by capsaicin. TRPV1 has traditionally been considered to cover a wide range of disease areas, including pain (inflammation, viscera, cancer and neuropathy, inflammatory bowel disease, interstitial cystitis, urinary incontinence, airway disease, pancreatitis and migraine), and its mechanisms include inflammation. Immunity, apoptosis, oxidative stress and so on. Recent studies have shown that TRPV1 is highly expressed not only in sensory neurons, but also in other functional brain regions, and is involved in the cellular process of neuronal injury. In addition, it was found that TRPV1 can reduce oxidative stress and related markers of cerebral infarction, and improve motor and cognitive function. It may play a role in the occurrence and development of neurodegenerative diseases. The purpose of this study was to investigate the neuroprotective effect of TRPV1 on Parkinson's disease rat model induced by 6-hydroxydopamine 6-OHDA-induced, and to provide a theoretical basis for its possible use as a new target for drug therapy. Methods one hundred and eight healthy male Wistar rats were randomly selected and their body weight was controlled at 220 g to 250 g, which was provided by the Animal experiment Center of Shandong University. They were randomly divided into normal control group (n = 20) and sham-operated group (n = 20 normal saline 6-OHDA group with right anterior medial bundle stereotactic injection (n = 40) treated with 6-OHDA capsaicin group (n = 40), the intervention group was treated by stereotactic injection of 6-OHDA capsaicin. After 6-OHDA injection, capsaicin 1mg/kg was injected intraperitoneally every day for 7 days, and the dose of capsaicin was 1 mg / kg ~ (-1) 路min ~ (-1) 路kg ~ (-1) 路min ~ (-1). Behavioral tests (including rotation test and open field test), Western blot detection of th and TRPV1 expression in substantia nigra and striatum, immunohistochemical detection of th neurons in substantia nigra and the content of MDA in rat brain were performed. All experimental data were processed with GraphPad Prism 5.0 software. The measurement data were described by the standard deviation of mean value (x 卤s), and the difference between the two groups was statistically significant by using t test to compare the mean value of the two groups. Results 1. The behavioral results showed that the number of induced rotation circles in the intervention group was significantly lower than that in the 6-OHDA group, and the walking distance and the average speed of the rats in the intervention group were significantly increased within one minute, and the residence time of the central lattice was significantly decreased, and the difference was statistically significant. Western blot showed that the expression of TRPV1 and th protein in substantia nigra and striatum in the intervention group was significantly higher than that in the other three groups, and the difference was statistically significant. The histochemical results showed that compared with 6-OHDA group, the number of th neurons in substantia nigra was significantly increased in the intervention group, in addition, the dendrites of th positive neurons and the length of axons in the substantia nigra were increased significantly in the intervention group. Compared with the 6-OHDA group, the levels of SOD in the right brain in the intervention group were significantly higher than those in the control group. The difference was statistically significant (P 0.05). There was no significant difference in the level of MDA between the left brain 6-OHDA group and the intervention group compared with the intervention group, and the difference was statistically significant (P 0.05) between the left brain 6-OHDA group and the intervention group, and there was no significant difference in the level of MDA between the left brain 6-OHDA group and the intervention group. Conclusion capsaicin can activate TRPV1 protein, regulate oxidative stress, protect dopaminergic neurons, improve motor function and relieve PD.
【学位授予单位】：山东大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R742.5;R-332

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