硫化氢对脑缺血再灌注损伤的保护作用及机制

发布时间：2018-03-16 21:16

本文选题：硫化氢　切入点：脑缺血再灌注　出处：《南华大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的：观察外源性硫化氢（hydrogen sulfide, H2S）对大鼠局灶性脑缺血再灌注损伤的保护作用并探讨其可能的机制。方法：50只SD雄性大鼠随机分为假手术组、缺血再灌注模型组、10μmol/kg、50μmol/kg和100μmol/kg外源性H2S供体硫氢化钠（sodiumhydrosulfide, NaHS）处理组，，每组10只。采用线栓法建立SD雄性大鼠缺血再灌注损伤模型，缺血2小时再灌注72小时。NaHS（10、50和100μmol/kg）在再灌注前30分钟腹腔注射；假手术组、缺血再灌注模型组给予等量生理盐水腹腔注射。再灌注6h、12h、24h、48h、72h对各组进行神经功能缺损评分。再灌注72小时后处死大鼠，采用2,3,5-氯化三苯基四氮唑（2,3,5-three phenyl tetrazolium chloride, TTC）染色观察并计算脑梗死的体积和比例。取缺血周边脑组织用分光光度法检测脑组织匀浆上清液中活性氧（reactive oxygen species, ROS）水平、丙二醛（malonaldehyde, MDA）含量和超氧化物歧化酶（superoxide dismutase, SOD）的活性；用双抗体夹心法测定脑组织匀浆上清液中总抗氧化力（total antioxidant capacity, T-AOC）的水平；用ELISA法测定脑组织匀浆上清液中还原型谷胱甘肽（reduced glutathione hormon,GSH）的水平。用PCR和Western blot检测缺血周边脑组织中脑源性神经营养因子（brain derived neurotrophic factor, BDNF）和酪氨酸激酶B（tyrosine kinase B,TrkB）mRNA和蛋白的表达。结果： 1、与缺血再灌注损伤模型组比较， NaHS（50和100μmol/Kg）处理组大鼠神经功能缺损明显改善，肌力明显增强，神经功能评分显著性降低（均P0.05）。 2、与假手术组比较，缺血再灌注损伤组大鼠缺血周边区脑组织中H2S浓度显著降低（P＜0.05），NaHS（50和100μmol/Kg）处理组大鼠缺血周边区脑组织中H2S浓度与模型组比较均显著升高，呈现剂量依赖性（均P0.05）。 3、与缺血再灌注损伤模型组比较，NaHS（50和100μmol/Kg）处理组的鼠脑梗死体积和比例显著降低，呈现剂量依赖性（均P0.05）。 4、与缺血再灌注损伤模型组比较，NaHS（50和100μmol/Kg）处理组大鼠缺血周边脑组织匀浆上清液中ROS水平和MDA含量显著降低（均P＜0.05），缺血周边脑组织匀浆上清液中T-AOC水平、SOD的活性和GSH水平显著增加（均P＜0.05）。 5、与缺血再灌注损伤模型组比较，NaHS（50和100μmol/Kg）处理组大鼠缺血周边脑组织中BDNF和TrkB的表达显著增加（均P㩳0.05）。结论：外源性H2S对脑缺血再灌注损伤具有保护作用，其机制可能与外源性H2S抑制氧化应激、上调BDNF和TrkB的表达有关。
[Abstract]:Aim: to observe the protective effect of exogenous hydrogen sulfide hydrogen sulfide (H _ 2S) on focal cerebral ischemia-reperfusion injury in rats and to explore its possible mechanism. Methods Fifty Sprague-Dawley male rats were randomly divided into sham-operation group, ischemia reperfusion model group (10 渭 mol / kg ~ (50) mol/kg) and exogenous H _ 2S donor sodium sulfide (NaHS) treatment group (n = 10). After 2 hours of ischemia and 72 hours of reperfusion, 50 and 100 渭 mol / kg NaHSN were injected intraperitoneally 30 minutes before reperfusion. The rats in the ischemia reperfusion model group were given intraperitoneal injection of the same amount of normal saline. The neurological impairment scores were evaluated in each group at 6 h, 12 h, 24 h, 48 h and 72 h after reperfusion. After 72 hours of reperfusion, the rats were killed. The volume and proportion of cerebral infarction were observed and calculated by TTC staining. The level of reactive oxygen species (Ros) in the supernatant of brain homogenate was detected by spectrophotometric method, and the content of reactive oxygen species (ROSs) in the supernatant of brain homogenate was determined by spectrophotometry. The content of malondialdehyde malonaldehydeand the activity of superoxide dismutase (SOD) were determined by double antibody sandwich method. The level of reduced glutathione reduced glutathione hormonium (GSH) in the supernatant of brain tissue homogenate was determined by ELISA method, and the expression of brain-derived neurotrophic factor (BDNFF) and tyrosine kinase (kinase) TrkBN mRNA and protein in peripheral ischemic brain tissue were detected by PCR and Western blot. Results:. 1.Compared with the model group of ischemia-reperfusion injury, NaHS(50 and 100 渭 mol / kg group significantly improved the nerve function defect, enhanced the muscle strength, and significantly decreased the neurological function score (all P 0.05). 2. Compared with the sham operation group, the concentration of H2S in the cerebral tissue of the ischemic peripheral area of rats in the ischemia-reperfusion group decreased significantly (P < 0.05), and the concentration of H _ 2S in the brain tissue of the ischemic peripheral area of the rats in the treatment group was significantly higher than that in the model group (P < 0.05), and the concentration of H _ 2S in the brain tissue of the ischemic peripheral area was significantly higher than that in the model group (all P < 0.05). 3Compared with the model group of ischemia-reperfusion injury, the volume and proportion of cerebral infarction in the NaHS 50 and 100 渭 mol / kg groups were significantly decreased in a dose-dependent manner (all P 0.05). 4. Compared with the model group of ischemia-reperfusion injury, the level of ROS and the content of MDA in the supernatant of cerebral homogenate of rats treated with NaHS 50 and 100 渭 mol / kg significantly decreased (all P < 0.05), the activity of T-AOC and the activity of T-AOC in the supernatant of cerebral homogenate of ischemic peripheral brain tissue decreased significantly (P < 0.05). The level of GSH increased significantly (all P < 0.05). 5. Compared with the model group of ischemia reperfusion injury, the expression of BDNF and TrkB in the ischemic peripheral brain tissue of rats treated with NaHS 50 and 100 渭 mol / kg were significantly increased (all P? 0.05. Conclusion: exogenous H2S has protective effect on cerebral ischemia-reperfusion injury, and its mechanism may be related to the inhibition of oxidative stress and up-regulation of BDNF and TrkB expression by exogenous H2S.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3

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