Necrostatin-1在小鼠脑出血中的作用及相关机制研究

发布时间：2018-05-06 11:32

本文选题：脑出血 + necrostatin-1　；参考：《苏州大学》2014年硕士论文

【摘要】：目的：在小鼠脑出血模型中，研究程序性坏死的特异性抑制剂necrostatin-1对脑出血引起的大脑损伤作用，探讨necrostatin-1对另外两种程序性细胞死亡方式（凋亡和自噬性细胞死亡）的影响。方法：采用胶原酶IV注入雄性ICR小鼠左侧纹状体建立脑出血（intracerebralhemorrhage, ICH）模型。在术前15min分别通过单侧侧脑室给药，并在伤后不同时间处死。首先利用组织缺损体积（lesion volume, LV）的方法评估necrostatin-1对ICH后组织损伤的作用；为了进一步验证脑组织的损伤是否与细胞损伤数相关，我们采用体视学显微镜观察并计数各组出血中心区以及周边区PI阳性细胞数。为了进一步探究necrostatin-1是否对其他两种程序性死亡方式（凋亡和自噬性细胞死亡）有影响，，我们利用免疫印记法分别检测了凋亡活性相关蛋白（Bcl-2，cleavedcaspase-3）和自噬活性相关蛋白（Beclin-1，LC3-II，p62）的表达水平。此外，为了进一步验证necrostatin-1对ICH后凋亡和自噬活性的影响，我们也联合利用凋亡的抑制剂zVAD及自噬抑制剂3-MA，检测了cleaved caspase-3和LC3-II的表达。结果：Necrostatin-1能够减少ICH引起的组织损伤体积和PI阳性细胞数；免疫印迹结果显示necrostatin-1能够降低ICH后自噬和凋亡相关蛋白的表达或活性。3-MA可以同时降低ICH后LC3-II和cleaved caspase-3的表达，而zVAD只降低了cleaved caspase-3表达却增加了LC3-II的表达。结论：在小鼠脑出血模型中，necrostatin-1可通过影响自噬和凋亡的相关蛋白表达或活性来发挥神经保护作用。
[Abstract]:Aim: to investigate the effects of necrostatin-1, a specific inhibitor of programmed necrosis, on brain injury induced by cerebral hemorrhage in mice, and to investigate the effects of necrostatin-1 on two other programmed cell death patterns (apoptosis and autophagic cell death). Methods: intracerebral hemorrhage (ICH) model was established by injecting collagenase IV into the left striatum of male ICR mice. 15min was administered through unilateral ventricle before operation, and was killed at different time after injury. The effect of necrostatin-1 on tissue damage after ICH was evaluated by tissue defect volume (LVV). The number of Pi positive cells in the central and peripheral areas of hemorrhage was observed and counted by stereological microscope. In order to further investigate the effects of necrostatin-1 on the other two programmed death patterns (apoptosis and autophagic cell death), we detected the expression levels of apoptosis-associated protein Bcl-2cleavedcaspase-3 and autophagy activity associated protein (Beclin-1) LC3-IIP62 by immunoblotting. In addition, in order to further verify the effect of necrostatin-1 on apoptosis and autophagy activity after ICH, we also detected the expression of cleaved caspase-3 and LC3-II by using zVAD, an inhibitor of apoptosis, and 3-MA. an inhibitor of autophagy. Results: Necrostatin-1 could reduce the volume of tissue injury and the number of Pi positive cells induced by ICH. Western blot showed that necrostatin-1 could decrease the expression of autophagy and apoptosis-related protein or the activity of apoptosis-related protein. 3-MA could decrease the expression of LC3-II and cleaved caspase-3 after ICH at the same time. ZVAD only decreased the expression of cleaved caspase-3, but increased the expression of LC3-II. Conclusion: necrostatin-1 may play a neuroprotective role by affecting the expression or activity of related proteins in autophagy and apoptosis in mice with intracerebral hemorrhage.
【学位授予单位】：苏州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743.34

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