糖尿病高血糖通过氧化损伤加重局灶性脑缺血再关注损伤

发布时间：2018-07-29 08:46

【摘要】：目的高血糖能够加重脑缺血再灌注损伤，但具体的途径和分子机制尚不清楚，本研究以高血糖局灶性脑缺血再灌注损伤大鼠作为模型，探讨高血糖加重脑缺血再灌注损伤中神经元及星形胶质细胞氧化损伤的作用。方法将雄性SD大鼠随机化分为正常血糖脑缺血再灌注组（简称正常血糖组）、糖尿病高血糖脑缺血再灌注组（简称糖尿病组）以及假手术对照组（简称假手术组），采用链脲佐菌素（Streptozotocin,STZ,55mg/kg）制备Ⅰ型糖尿病大鼠模型，糖尿病高血糖形成后3d内实施MCAO缺血30min，再灌注1d，3d，7d，14d。分别进行组织学、8羟基脱氧鸟苷（8-hydroxy-2’deoxyguanosine，8-OHdG）免疫组织化学、NeuN和8-OHdG免疫荧光双标记及GFAP和8-OHdG免疫荧光双标记技术，对比观察各组神经元及星形胶质细胞的损伤及其与8-OHdG的关系。结果：组织学显示，正常血糖组再灌注1d脑组织出现水肿，糖尿病组较正常血糖组有所加重，出现较多的固缩神经元，再灌注3d脑组织水肿进一步加重，固缩神经元进一步增多,可见多数细胞溶解坏死，存留泡沫样结构，同样，糖尿病组表现更为严重；再灌注7d正常血糖组神经元固缩和脑水肿明显减轻，糖尿病组仍可见少数神经元固缩和脑水肿；再灌注14d正常血糖组神经元固缩和脑水肿消失，胶质细胞增加，糖尿病组可见轻度脑水肿。免疫组化显示，再灌注3d，正常血糖组及糖尿病组8-OHdG阳性细胞数量均明显增加，高血糖组8-OHdG阳性细胞数量明显高于正常血糖组（P0.05），再灌注7d和14d8-OHdG免疫阳性细胞明显减少，但仍多于假手术组（P0.05）；免疫荧光双标记提示，，再灌注3d时，正常血糖组及糖尿病组8-OHdG阳性的神经元均达高峰，且糖尿病组的8-OHdG阳性神经元明显高于正常血糖组（P0.05），再灌注7d和14d时，8-OHdG阳性的神经元有所减少；且正常血糖组再灌注3d8-OHdG阳性的星形胶质细胞开始增加，再灌注7d时达高峰，糖尿病组较正常血糖组双标阳性细胞明显增加（P0.05），再灌注14d双标阳性细胞数有所下降,但仍多于假手术组。结论：（1）糖尿病高血糖可以加重脑缺血再灌注损伤，使局灶性脑缺血梗死区及其周围脑组织水肿加重，梗死区扩大，神经元变性和凋亡增加，恢复时间延长；（2）脑缺血再灌注引起中枢神经系统氧化损伤，局灶性脑缺血30min，再灌注1天氧化损伤明显增加，再灌注3天时达到高峰，再灌注7天氧化损伤明显减少；糖尿病高血糖能够明显加重再灌注1天和3天时的氧化损伤，再灌注7天氧化损伤减轻的幅度明显大于正常血糖组；（3）局灶性脑缺血30min，再灌注1天和3天氧化损伤明显增加，氧化损伤主要发生在神经元；糖尿病高血糖能够明显加重神经元的氧化损伤，星形胶质细胞氧化损伤较轻微。
[Abstract]:Objective hyperglycemia can aggravate cerebral ischemia-reperfusion injury, but the specific pathway and molecular mechanism are not clear. In this study, focal cerebral ischemia-reperfusion injury induced by hyperglycemia in rats was used as model. To investigate the effect of hyperglycemia on oxidative injury of neurons and astrocytes in cerebral ischemia reperfusion injury. Methods male Sprague-Dawley rats were randomly divided into normal blood glucose cerebral ischemia-reperfusion group, diabetic hyperglycemic cerebral ischemia reperfusion group and sham operation control group. The rat model of type 1 diabetes mellitus was established by using streptozotocin (STZ) 55 mg / kg. Diabetic hyperglycemia was performed within 3 days after MCAO ischemia for 30 minutes and reperfusion for 1 day for 3 days and 7 days for 14 days. Immunohistochemical staining of 8-hydroxy-2-deoxyguanosine 8-OHdG (Neun and 8-OHdG) and double labeling of GFAP and 8-OHdG were performed respectively. The damage of neurons and astrocytes and their relationship with 8-OHdG were observed. Results: histologically, there were edema in brain tissue of normal blood glucose group on the 1st day after reperfusion, more pyknotic neurons appeared in diabetes group than in normal blood glucose group, and the edema of brain tissue was further aggravated at 3 days after reperfusion. The pyknotic neurons increased further, most of the cells dissolved and necrotized, and the foam structure remained. Similarly, the diabetic group showed more serious symptoms, while the normal blood glucose group on the 7th day after reperfusion significantly alleviated the pyknosis and brain edema. There were still a few neurons pyknosis and brain edema in the diabetic group, while the neuron pyknosis and brain edema disappeared in the normal blood glucose group on the 14th day after reperfusion, the glial cells increased, and mild brain edema was observed in the diabetic group. Immunohistochemistry showed that the number of 8-OHdG positive cells in normal blood glucose group and diabetic group was significantly increased on the 3rd day after reperfusion, and the number of 8-OHdG positive cells in hyperglycemia group was significantly higher than that in normal blood glucose group (P0.05), and the number of 14d8-OHdG immunoreactive cells and 14d8-OHdG immunoreactive cells in hyperglycemia group decreased significantly at 7 days after reperfusion. But it was still more than sham operation group (P0.05), immunofluorescence double labeling showed that the 8-OHdG positive neurons in normal blood glucose group and diabetes group reached the peak at 3 days after reperfusion. The 8-OHdG positive neurons in the diabetic group were significantly higher than those in the normal blood glucose group (P0.05), and the number of 8-OHdG positive neurons decreased on the 7th and 14th days after reperfusion, and the number of 3d8-OHdG positive astrocytes in the normal blood glucose group began to increase and reached the peak at the 7th day after reperfusion. The number of double labeled positive cells in diabetes group was significantly higher than that in normal blood glucose group (P0.05), and the number of double labeled positive cells decreased on the 14th day after reperfusion, but still more than that in sham operation group. Conclusion: (1) Diabetic hyperglycemia can aggravate cerebral ischemia-reperfusion injury, aggravate edema, enlarge infarct area, increase neuronal degeneration and apoptosis, and prolong recovery time; (2) oxidative injury of central nervous system was induced by cerebral ischemia-reperfusion. After 30 min of focal cerebral ischemia, the oxidative injury increased significantly on 1 day after reperfusion, reached the peak at 3 days after reperfusion, and decreased significantly on 7 days after reperfusion. Diabetic hyperglycemia could significantly aggravate oxidative injury on 1 and 3 days after reperfusion, and the extent of oxidative damage at 7 days after reperfusion was significantly higher than that in normal blood glucose group, (3) oxidative damage was significantly increased at 30 min after focal cerebral ischemia, 1 day and 3 days after reperfusion. The oxidative damage occurred mainly in neurons, and diabetic hyperglycemia significantly aggravated the oxidative damage of neurons, while the oxidative damage of astrocytes was slight.
【学位授予单位】：宁夏医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.3

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