基于痛性相关神经肽CGRP、SP及其受体NK-1探讨电针防治急性发作期偏头痛机制

发布时间：2018-06-29 17:21

本文选题：电针 + 偏头痛　；参考：《首都医科大学》2017年硕士论文

【摘要】：目的偏头痛是常见的原发性头痛,多以单侧或双侧颞部反复疼痛为主要的临床表现。针刺在治疗偏头痛的临床疗效确切,但其治疗机制尚未完全明确。随着医学的不断进步,对偏头痛发病机制研究从细胞水平逐渐深入到神经肽水平,大量研究[1,2]发现多种痛性神经肽与偏头痛的发作密切相关,如神经肽家族群SP及其受体NK-1、降钙素相关肽(CGRP)、5-羟色胺(5-HT)、一氧化氮(NO)、内皮素等,其中P物质与降钙素相关肽(CGRP)作为神经递质在感觉传导和痛觉调制中作用越来越引起研究者的注意。有大量实验[3,4]已证明神经肽SP及CGRP在神经调制中既能传递痛觉信息,有致痛作用;又能促进脑啡肽(enkehalin,ENK)的释放,从而达到镇痛作用。故本实验通过皮下注射硝酸甘油备制模型,观察电针对急性发作期偏头痛大鼠中脑SP、NK-1分布与表达及血浆CGRP浓度变化的影响,来探讨电针防治急性发作期偏头痛相关机制。方法250±50g健康Wistar雄性大鼠80只,每4只一个笼子,清洁级实验室喂养,自由摄食、饮水。饲养温度(22±1?C),湿度40%~70%,噪音60分贝,换气次数10~20次/小时,光照时间每天12小时(8:00~20:00),光照时间模拟昼夜交替,适应性饲养7天。随机分为4组:空白组、模型组、电针组、伪电针组,每组20只。空白组不作任何处理,正常饮食摄水饲养7天,于第8天皮下注射生理盐水;模型组不作电针干预治疗,正常饮食摄水饲养7天,于第8天皮下注射硝酸甘油制备偏头痛大鼠模型;电针组通过电针双侧太阳、风池、太冲、足临泣穴干预7天;伪电针组进行非穴位(非穴位点为非经非穴部位,其位置不固定)电针干预7天后,于第8天分别对两组进行皮下注射硝酸甘油制备偏头痛大鼠模型。造模成功后,对两组进行相应的电针干预治疗。本实验主要采用以下几项作为偏头痛大鼠观察指标:挠头、爬笼、咬尾、往返运动的次数并计数(每个症状出现1次计1分);采用酶联免疫吸附法(ELISA)测定大鼠血浆中CGRP的浓度;免疫组化检测中脑导水管周围灰质区SP、NK-1阳性细胞数;通过实时荧光定量PCR技术检测大鼠中脑导水管周围灰质区SP及其受体NK-1基因表达。结果1、电针对偏头痛大鼠血浆CGRP浓度变化的影响:与空白组比较,模型组、伪电针组血浆CGRP浓度均明显升高(P0.05),电针组无明显变化(P0.05);与模型组比较,电针组浓度明显降低(P0.05),伪电针组无明显变化(P0.05);与伪电针组比较,电针组浓度明显降低(P0.05)。提示电针能显著降低急性发作期偏头痛大鼠血浆中CGRP浓度,其疗效明显优于伪电针治疗。2、电针对大鼠中脑组织SP及NK-1基因表达影响:与空白组比较,模型组、电针组、伪电针组SP、NK-1m RNA表达量明显增高(P0.05);与模型组比较,电针组、伪电针组SP、NK-1m RNA表达量明显降低(P0.05);与伪电针组比较,电针组SP、NK-1 m RNA表达量明显降低(P0.05)。提示电针能显著降低急性发作期偏头痛PAG区的SP及NK-1的基因表达水平,其疗效明显优于伪电针治疗。3、电针对大鼠中脑导水管周围灰质区SP及NK-1蛋白表达及分布的影响。SP阳性细胞数:与空白组(A组)比较,模型组(B组)、伪电针组(D组)阳性细胞数均明显升高(P0.05),电针组(C组)无明显变化(P0.05);与模型组(B组)比较,电针组(C组)、伪电针组(D组)阳性细胞数均明显降低(P0.05);与伪电针组(D组)相比较,电针组(C组)阳性细胞数明显降低(P0.05)。NK-1阳性细胞数表达:与空白组(A组)比较,模型组(B组)、电针组(C组)、伪电针组(D组)均明显升高,且差异有统计学意义(P0.05);模型组(B组)、电针组(C组)、伪电针组(D组)三组间各自比较均无明显差异(P0.05)。提示,电针能显著降低急性发作期偏头痛大鼠中脑SP的阳性分布及表达,其疗效明显优于伪电针治疗;对于NK-1阳性表达及分布的影响有降低趋势,但无明显变化。结论电针通过下调急性发作期偏头痛大鼠痛性相关神经肽CGRP、SP表达量,发挥防治偏头痛的作用。
[Abstract]:Objective migraine is a common primary headache, mostly with unilateral or bilateral temporal and recurrent pain as the main clinical manifestation. Acupuncture is effective in the treatment of migraine, but its mechanism of treatment is not completely clear. With the continuous progress of medicine, the study of the pathogenesis of migraine from cell level to neuropeptide level, [1,2] found that a variety of painful neuropeptides are closely related to the attack of migraine, such as the neuropeptide family group SP and its receptor NK-1, calcitonin related peptide (CGRP), 5- serotonin (5-HT), nitric oxide (NO), endothelin, etc., in which P substance and calcitonin related peptide (CGRP) Act more and more as neurotransmitters in sensory conduction and pain modulation Researchers' attention. A large number of experiments [3,4] have proved that neuropeptide SP and CGRP can not only transmit pain information, cause pain, but also promote the release of enkephalin (enkehalin, ENK) and thus achieve analgesic effect. Therefore, the experiment was made by subcutaneous injection of nitroglycerin to observe the effect of Electroacupuncture on acute migraine rats. The influence of the distribution and expression of SP, NK-1 and the change of CGRP concentration in the plasma to discuss the mechanism of electroacupuncture in the prevention and treatment of acute migraine. Methods 80 healthy Wistar male rats of 250 + 50g, every 4 cages, free feeding, drinking water, feeding temperature (22 + 1? C), humidity 40%~70%, noise 60 decibels, air exchange times 10~20 The light time was 12 hours a day (8:00~20:00), and the light time was simulated day and night and adapted for 7 days. It was randomly divided into 4 groups: blank group, model group, electroacupuncture group, pseudo-needle group, 20 in each group. The blank group was not treated with any treatment, normal diet for 7 days and eighth days by subcutaneous injection of saline; the model group did not do electroacupuncture intervention, The rat model of migraine was prepared by subcutaneous injection of nitroglycerin on the eighth day for 7 days, and the electroacupuncture group was treated with Electroacupuncture of the sun, the wind pond, the Tai Chi, and the foot and the weeping point for 7 days. The pseudo electroacupuncture group carried out the non acupoint (non acupoint points for non acupoint points, and its position was not fixed) for 7 days, and the two groups were carried out on the eighth day respectively. The rat model of migraine was prepared by injection of nitroglycerin. After the model was successful, the two groups were treated with electroacupuncture. This experiment mainly used the following indexes as observation indexes of migraine rats: bending head, crawling cage, tail biting, and counting the number of round-trip times (1 times each symptom was 1); enzyme linked immunosorbent assay (ELISA) The concentration of CGRP in the rat plasma; immunohistochemistry to detect the number of SP, NK-1 positive cells in the periaqueductal gray area of the middle cerebral aqueduct; the expression of SP and its receptor NK-1 gene in the periaqueductal gray area of the rat by real-time fluorescence quantitative PCR. Results 1, the effect of Electroacupuncture on the plasma CGRP concentration in the migraine rats: compared with the blank group, the model group, The concentration of plasma CGRP in the pseudo electroacupuncture group increased significantly (P0.05), and there was no obvious change in the electroacupuncture group (P0.05). Compared with the model group, the concentration of electroacupuncture group decreased significantly (P0.05), and there was no obvious change in the pseudo-electroacupuncture group (P0.05), and the concentration of electroacupuncture group decreased significantly (P0.05) compared with the pseudo electroacupuncture group (P0.05). It suggested that electroacupuncture could significantly reduce the CGRP in the plasma of migraine rats at acute attack. The effect was obviously better than that of the pseudo-electroacupuncture treatment of.2, and the effect of Electroacupuncture on the expression of SP and NK-1 gene in the middle brain tissue of rats: compared with the blank group, the expression of SP and NK-1m RNA in the model group, the electroacupuncture group and the pseudo electroacupuncture group increased significantly (P0.05). Compared with the model group, the expression of SP and NK-1m RNA in the electroacupuncture group, the pseudo-needle group and the NK-1m RNA was obviously reduced (P0.05); compared with the pseudo electroacupuncture group, The expression of SP and NK-1 m RNA decreased significantly in the electroacupuncture group (P0.05). It was suggested that electroacupuncture could significantly reduce the gene expression level of SP and NK-1 in the PAG region of acute migraine, and the effect was obviously better than that of the pseudo-electroacupuncture in the treatment of.3. The number of SP and NK-1 protein in the periaqueductal gray area of the rat middle cerebral aqueduct affected the number of.SP positive cells and the blank group. Compared with the model group (group B), the number of positive cells in the pseudo electroacupuncture group (group D) increased significantly (P0.05), and there was no significant change in the electroacupuncture group (group C) (P0.05). Compared with the model group (group B), the number of positive cells in the electroacupuncture group (group C) and the pseudo electroacupuncture group (D group) decreased significantly (P0.05), and the number of positive cells in the electroacupuncture group (C group) was significantly decreased (P0.05) compared with the pseudo electroacupuncture group (D group). The expression of NK-1 positive cells: compared with the blank group (group A), the model group (group B), the electroacupuncture group (group C), the pseudo electroacupuncture group (D group) were all significantly increased, and the difference was statistically significant (P0.05). There was no significant difference between the three groups in the model group (B group), the electroacupuncture group (C group) and the pseudo electroacupuncture group (D group). It suggested that the electroacupuncture could significantly reduce the acute attack stage head. The positive distribution and expression of SP in the midbrain of the rats were significantly better than that of the pseudo-electroacupuncture treatment, and the effect on the positive expression and distribution of NK-1 had a decreasing trend, but there was no obvious change. Conclusion electroacupuncture can play a role in the prevention of migraine by reducing the pain related neuropeptide CGRP and SP expression in the acute attack stage of migraine rats.
【学位授予单位】：首都医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R245

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