头穴透刺预处理对脑缺血再灌注大鼠内源性神经干细胞的影响

发布时间：2018-07-01 19:18

本文选题：头穴透刺 + 预处理　；参考：《黑龙江省中医药科学院》2016年硕士论文

【摘要】：目的:根据中医“治未病”思想,研究探讨头穴透刺预处理对大鼠脑缺血再灌注后内源性NSCs的增殖、分化以及对BDNF、b FGF的影响,明确头穴透刺预处理对脑缺血性卒中的预防与治疗作用,从干细胞这一角度,探讨头穴透刺预处理预防缺血性脑卒中的可能机制。方法:实验选取雄性Wistar大鼠120只,采用随机数字表方法将大鼠随机分为:假手术组、模型组、实验组,每组各40只,每组又分为3、7、14、21d四个时间点,每个时间点10只;采用改良线栓法制备大鼠脑缺血再灌注模型(MCAO),实验组于造模前连续针刺7d,针刺选取百会透双侧太阳穴,每次针刺治疗时间为20分钟,针刺治疗后造模,模型组仅造模,不做其他处理,假手术组除不插线外,余操作同模型组;通过腹腔注射Brdu来标记增殖的细胞,分别于3、7、14、21d四个时间点取材来对实验对象进行观察研究。采用神经症状学评分确定模型成功标准,通过TTC染色和HE染色来了解脑缺血再灌注后脑组织的病理学改变。通过免疫组化观察Brdu、Nestin阳性细胞,确定NSCs的增殖情况,通过免疫荧光化学技术观察Brd U/GFAP、Brd U/NSE双标细胞,确定NSCs分化情况。免疫组化法观察BDNF、b FGF变化,确定对NSCs增殖的可能机制。结果:1.实验组与模型组各个时间点比较神经功能缺损评分均降低,实验组在7、14、21d点神经功能评分与模型组比较具有显著性差异(P0.05)。2.实验组各个时间点与模型组比较Brd U免疫阳性细胞表达均增强,并随时间变化,3d时开始增加,7d时达到高峰,14d开始下降,各个时间点阳性细胞数量与模型组相比,有显著性差异(P0.05)。3.实验组Nestin免疫阳性细胞表达明显增强,3d时开始增加,7d时达到高峰,7、14、21d与模型组阳性细胞数相比,有显著性差异(P0.05)。4.实验组Brd U/GFAP、Brd U/NSE免疫荧光双标阳性细胞明显增加,于缺血再灌注后7d达高峰,14d开始下降,与模型组7、14d比较有显著性差异(P0.05)。5.实验组BDNF的表达明显增强,于缺血再灌注21d时达高峰,与模型组7、14、21d相比有显著性差异(P0.05)。6.实验组b FGF的表达明显增强,于7d时达高峰。与模型组各个时间点比较有显著性差异(P0.05)。结论:1、头穴透刺预处理能显著降低脑缺血再灌注后大鼠神经功能症状评分,促进其神经功能恢复。2、头穴透刺预处理能显著促进脑缺血再灌注后大鼠脑组织Brdu、Nestin、Brd U/GFAP、Brd U/NSE表达增加,从而促进NSCs的增殖和分化。3、头穴透刺预处理能促进脑缺血再灌注后大鼠神经营养因子b FGF、BDNF的表达,从而有助于神经功能的恢复。
[Abstract]:Objective: to investigate the effects of scalp acupoint penetration preconditioning on the proliferation and differentiation of endogenous NSCs after cerebral ischemia-reperfusion in rats and the effects on BDNF bFGF according to the idea of "treating disease without disease" in traditional Chinese medicine (TCM). To clarify the preventive and therapeutic effects of scalp acupoint penetration preconditioning on ischemic stroke and to explore the possible mechanism of scalp acupoint penetration preconditioning in preventing ischemic stroke from the point of view of stem cells. Methods: 120 male Wistar rats were selected and randomly divided into four groups: sham operation group, model group, experimental group, 40 rats in each group. The rat model of cerebral ischemia-reperfusion (MCAO) was established by modified thread embolization method. The experimental group was treated continuously for 7 days before the model was made. The acupuncture group was treated with Baihui dialysed bilateral temples for 20 minutes each time. The model was made after acupuncture treatment, and only the model was made in the model group. Without any other treatment, the sham-operation group operated with the same model group except without wire insertion, and the proliferating cells were labeled by intraperitoneal injection of Brdu. The successful criteria of the model were determined by neurosymptomatic score, and the pathological changes of brain tissue after cerebral ischemia and reperfusion were studied by TTC staining and HE staining. The proliferation of NSCs was determined by immunohistochemical observation of Brdun nestin positive cells, and the differentiation of NSCs was determined by using immunofluorescence technique to observe BrdU / GFAP / Brd / NSE double labeled cells. The changes of BDNFB FGF were observed by immunohistochemical method to determine the possible mechanism of proliferation of NSCs. The result is 1: 1. Compared with the model group, the neurological deficit score of the experimental group and the model group decreased at each time point, and there was significant difference between the experimental group and the model group on the 21st day (P0.05). 2. Compared with the model group, the expression of BrdU immunoreactive cells in the experimental group increased at each time point, and increased at the 3rd day and reached the peak at the 7th day. The number of the positive cells in each time point was significantly different from that in the model group (P0.05). 3. The expression of BrdU immunoreactive cells in the experimental group was significantly higher than that in the model group (P0.05). The expression of nestin immunoreactive cells in the experimental group was significantly higher than that in the model group (P0.05). BrdU / GFAPG BrdU / NSE double immunofluorescence positive cells increased significantly in the experimental group, and began to decrease on the 7th day after ischemia and reperfusion, reaching the peak at the 14th day, which was significantly different from that in the model group (P0.05) on the 14th day (P0.05). The expression of BDNF in the experimental group was significantly increased and reached its peak at 21 days after ischemia reperfusion, which was significantly different from that in the model group on the 21st day (P0.05). The expression of bFGF in the experimental group was significantly increased and reached its peak at 7 days. There was significant difference between the model group and each time point (P0.05). Conclusion the scalp point penetration preconditioning can significantly decrease the neurological function symptom score and promote the recovery of nerve function after cerebral ischemia reperfusion in rats. The scalp penetration acupuncture preconditioning can significantly increase the expression of Brd Nestin + Brd / GFAPP + Brd / NSE in brain tissue after cerebral ischemia and reperfusion. In order to promote the proliferation and differentiation of NSCs, scalp acupoint penetration preconditioning can promote the expression of neurotrophic factor b FGFN BDNF after cerebral ischemia-reperfusion in rats, thus contributing to the recovery of nerve function.
【学位授予单位】：黑龙江省中医药科学院
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R245

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