人参皂苷Rb1对慢性缺血损伤大鼠脑组织中自噬表达的影响
发布时间：2018-11-01 18:16
【摘要】：目的:探讨人参皂苷Rb1对自噬调节的作用,以期能寻找一种减轻脑缺血缺氧后脑组织损伤的中药,进而减轻脑缺血缺氧对机体带来的影响。方法:将30只SD大鼠随机分成6组,每组5只,分别为高剂量组、中剂量组、低剂量组、空白对照组、假手术组、未做处理组,通过永久性结扎双侧颈总动脉模拟脑缺血缺氧模型,分别按照1Omg/kg、5mg/kg、2.5mg/kg的浓度在造模后1-28天对高剂量组、中剂量组、低剂量组进行腹腔注射,每天一次,并对假手术组和空白对照组每天给予超纯水0.5ml,未做处理组不进行腹腔注射作为对照。同时1、分别在造模前、造模45天后、造模90天后对各组大鼠进行为期10天的水迷宫实验(具体包括定位航行实验、工作记忆实验)以通过观察各组大鼠寻找平台的功能来评估大鼠的学习记忆能力进而评价其认知功能。2、在最后一次水迷宫实验结束后对大鼠断头取脑,收集其额叶及海马组织,①Western blot检测大鼠脑组织里Caspase-3、LC3B的含量,从而了解大鼠脑组织的自噬水平。②将脑组织HE染色后在光学显微镜下观察(1)脑:有没有变性、坏死的皮质神经元,有没有增生的胶质细胞,髓质有没有出现水肿。(2)海马:海马各区域的椎体细胞有没有变性及坏死等改变。同时对脑组织出现的各种以上改变进行程度轻重的评分。结果:①与正常对照组、假手术组初次水迷宫后即获得学习记忆能力,在第45天及90天后寻找平台时间明显缩短相比,模型组大鼠出现明显认知功能障碍,具体表现为不管是工作记忆还是定位航行,模型组大鼠寻找平台的持续时间及潜伏期均延长(P0.05),上台前搜索距离明显延长(P0.001),这一表现在造模90天后表现更显著。而人参皂苷Rb1可降低脑缺血缺氧大鼠寻找平台的持续时间及潜伏期(P0.05),同时明显缩短模型大鼠上台前的搜索距离(P0.001),浓度越高效果越明显。②所有脑缺血缺氧模型组的Caspase-3含量增高(P0.05),LC3B含量降低(P0.05),人参皂苷Rb1作用于模型鼠后可抑制Caspase-3的表达同时促进LC3B的表达,且浓度越高效果越明显。③HE染色后观察模型组大鼠脑组织后可见:脑组织结构与未做处理组相比,大多数区域皮层结构消失,神经元坏死,局部组织因高度疏松而表现为网状,可以看到血管及胶质细胞增殖;1区及2区海马结构不清楚,大多数锥体细胞坏死、消失,残留的个别坏死细胞结构不明;3区大多锥体细胞形状不规则、核固缩、体积变性;病灶改变程度评分显示模型组大鼠脑组织出现极重度改变;高剂量组、中剂量组脑组织为轻度改变,低剂量组脑组织为中度改变,空白剂量组脑组织为极重度改变,假手术组脑组织为轻微改变。表明人参皂苷Rb1可改善模型组大鼠的脑组织坏死能力且浓度越高改善作用越明显。结论:人参皂苷Rb1可通过改善脑缺血缺氧模型组大鼠的学习记忆能力、调节自噬因子的表达、减轻脑组织坏死等作用降低脑缺血缺氧大鼠的脑组织损伤程度。
[Abstract]:Objective: To explore the effect of ginseng soap Rb1 on autophagy, in order to find a traditional Chinese medicine for alleviating brain injury after cerebral ischemia and anoxia, and to reduce the effects of cerebral ischemia and hypoxia on organism. Methods: Thirty SD rats were randomly divided into 6 groups, 5 rats in each group, the high dose group, the middle dose group, the low dose group, the blank control group, the sham operation group and the untreated group. 2. The concentration of 5mg/ kg was injected intraperitoneally in high dose group, medium dosage group and low dose group for 1 to 28 days after the model molding, and 0.5ml of ultrapure water was administered to the sham operation group and the blank control group every day, and no abdominal cavity injection was performed in the untreated group as the control group. At the same time, after the model was established for 45 days, a 10-day water maze test was carried out on each group of rats (including positioning navigation experiment) after 90 days respectively. working memory experiment) to evaluate the learning and memory ability of rats by observing the function of each group of rat searching platform to evaluate its cognitive function. Western blot was used to detect the level of Caspase-3 and LC3B in brain tissue of rats. He stained the brain tissue with HE and observed (1) the brain under an optical microscope: whether there was degeneration, necrosis of the cortical neurons, the presence of proliferating glial cells, and the presence of edema in the medulla. (2) hippocampus: there is no change in the degeneration and necrosis of the vertebral body cells in each region of the hippocampus. At the same time, the severity of the changes in brain tissue was scored. Results: Compared with the normal control group and the sham operation group, after the initial water maze, the learning and memory ability was acquired. After 45 days and 90 days, there was obvious cognitive dysfunction in the model group. The duration and latency of search platform for model group were prolonged (P0.05). The duration and incubation period (P <0.01) of the model rats were significantly shortened (P <0.01), and the higher the concentration was, the higher the concentration was. The expression of Caspase-3 was decreased (P0.05). The expression of Caspase-3 was inhibited at the same time, and the higher the concentration was. After HE staining, it was found that the structure of the brain tissue disappeared, the neurons were necrotic, the local tissue was reticulate because of the high degree of osteoporosis, and the proliferation of vascular and glial cells could be seen. In 1 and 2 regions, the structure of hippocampal formation was not clear, most of the pyramidal cells were necrotic, disappeared, and the residual individual necrotic cells were unknown. In the high-dose group, the brain tissue of the middle-dose group was slightly changed, the brain tissue of the low-dose group was moderately changed, the brain tissue of the blank dose group was very severe, and the brain tissue of the sham-operated group was slightly changed. The results showed that Rb1 could improve the brain tissue necrosis and the higher the concentration. Conclusion: Ginseng soap Rb1 can improve the learning and memory ability of rats with cerebral ischemia and hypoxia model, regulate the expression of autophagy factor, reduce the brain tissue necrosis, and decrease the damage degree of brain tissue in rats with cerebral ischemia and hypoxia.
【学位授予单位】：南京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285.5
[Abstract]:Objective: To explore the effect of ginseng soap Rb1 on autophagy, in order to find a traditional Chinese medicine for alleviating brain injury after cerebral ischemia and anoxia, and to reduce the effects of cerebral ischemia and hypoxia on organism. Methods: Thirty SD rats were randomly divided into 6 groups, 5 rats in each group, the high dose group, the middle dose group, the low dose group, the blank control group, the sham operation group and the untreated group. 2. The concentration of 5mg/ kg was injected intraperitoneally in high dose group, medium dosage group and low dose group for 1 to 28 days after the model molding, and 0.5ml of ultrapure water was administered to the sham operation group and the blank control group every day, and no abdominal cavity injection was performed in the untreated group as the control group. At the same time, after the model was established for 45 days, a 10-day water maze test was carried out on each group of rats (including positioning navigation experiment) after 90 days respectively. working memory experiment) to evaluate the learning and memory ability of rats by observing the function of each group of rat searching platform to evaluate its cognitive function. Western blot was used to detect the level of Caspase-3 and LC3B in brain tissue of rats. He stained the brain tissue with HE and observed (1) the brain under an optical microscope: whether there was degeneration, necrosis of the cortical neurons, the presence of proliferating glial cells, and the presence of edema in the medulla. (2) hippocampus: there is no change in the degeneration and necrosis of the vertebral body cells in each region of the hippocampus. At the same time, the severity of the changes in brain tissue was scored. Results: Compared with the normal control group and the sham operation group, after the initial water maze, the learning and memory ability was acquired. After 45 days and 90 days, there was obvious cognitive dysfunction in the model group. The duration and latency of search platform for model group were prolonged (P0.05). The duration and incubation period (P <0.01) of the model rats were significantly shortened (P <0.01), and the higher the concentration was, the higher the concentration was. The expression of Caspase-3 was decreased (P0.05). The expression of Caspase-3 was inhibited at the same time, and the higher the concentration was. After HE staining, it was found that the structure of the brain tissue disappeared, the neurons were necrotic, the local tissue was reticulate because of the high degree of osteoporosis, and the proliferation of vascular and glial cells could be seen. In 1 and 2 regions, the structure of hippocampal formation was not clear, most of the pyramidal cells were necrotic, disappeared, and the residual individual necrotic cells were unknown. In the high-dose group, the brain tissue of the middle-dose group was slightly changed, the brain tissue of the low-dose group was moderately changed, the brain tissue of the blank dose group was very severe, and the brain tissue of the sham-operated group was slightly changed. The results showed that Rb1 could improve the brain tissue necrosis and the higher the concentration. Conclusion: Ginseng soap Rb1 can improve the learning and memory ability of rats with cerebral ischemia and hypoxia model, regulate the expression of autophagy factor, reduce the brain tissue necrosis, and decrease the damage degree of brain tissue in rats with cerebral ischemia and hypoxia.
【学位授予单位】：南京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285.5
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