Basic Investigations in Brain Remodeling

用选择性5型磷酸二酯酶（PDE 5）抑制剂如西地那非治疗中风， 显著促进功能恢复，同时显著增加神经发生， 当在中风发作后1或7天开始治疗时，血管生成减少。在本申请中，我们寻求 阐明神经发生，神经母细胞迁移和耦合的基本机制， 脑卒中后有无西地那非治疗的神经发生和血管生成 神经修复剂我们的假设是西地那非介导了脑卒中患者cGMP水平的升高 脑：1）通过缩短细胞周期的长度来促进神经祖细胞的增殖， 结果从细胞周期蛋白依赖性激酶抑制剂的表达减少; 2）增强神经母细胞 通过增加基质衍生因子-la的表达向缺血边界区域迁移 （SDF-1a）、CXCR4和基质金属蛋白酶（MMPs）; 3）促进血管生成， 允许的小生境，以促进神经发生并引导成神经细胞向缺血的 4）PI3K/Akt信号通路介导西地那非增强的神经发生， 成神经细胞迁移。所提出的实验旨在验证这些假设。我们将 研究西地那非是否放大细胞周期动力学和细胞分裂的方向， 脑室下区神经前体细胞和干细胞的增殖和分化。使用 逆转录病毒载体，siRNA和延时显微镜，我们将深入研究的分子机制，例如。 SDF-1/CXCR4和MMPs，它们促进了缺血脑中的神经母细胞迁移， 西地那非观察西地那非诱导的血管生成对神经发生和成神经细胞的影响 迁移，我们将测量血管生成和神经发生以及分泌的蛋白质之间的相互作用 被内皮细胞所控制。通过阻断SDF-1a和MMPs，我们将检查SDF-1a和MMPs是否 由内皮细胞分泌介导成神经细胞迁移。使用特定的抑制剂，我们将研究 阻断PI3K/Akt信号通路是否减弱西地那非对神经发生的作用， 成神经细胞迁移和血管生成。这些基础科学研究将阐明 中风后大脑重塑，以及cGMP如何在成人大脑中放大这一过程，这可能 从而产生了促进中风恢复期间脑重塑的新方法。

Treatment of stroke with a selective inhibitor of phosphodiesterase type 5 (PDE5), e.g.sildenafil, significantly promotes functional recovery concomitant with significant increases of neurogenesis and angiogenesis when the treatment is initiated 1 or 7 days after stroke onset. In this application, we seek to elucidate fundamental mechanisms of neurogenesis, neuroblast migration and the coupling of neurogenesis and angiogenesis after stroke with and without treatment with sildenafil as the neurorestorative agent. Our hypotheses are that sildenafil mediated increases of cGMP levels in stroke brain: 1) promote proliferation of neural progenitor cells via shortening the length of the cell cycle which results from a decreased expression of cyclin-dependent kinase inhibitors; 2) enhance neuroblast migration towards the ischemic boundary regions via increasing expression of stromal-derived factor-la (SDF-1a), CXCR4 and matrix metalloproteinases (MMPs); 3) foster angiogenesis which generates a permissive niche to promote neurogenesis and to guide neuroblast migration towards the ischemic boundary regions; and 4) the PI3K/Akt signaling pathway mediates sildenafil-enhanced neurogenesis and neuroblast migration. The proposed experiments have been designed to test these hypotheses. We will investigate whether sildenafil amplifies cell cycle kinetics and the orientation of cell division that regulate the proliferation and differentiation of neural progenitor and stem cells in the subventricular zone. Using retroviral vector, siRNA, and time-lapse microscopy, we will delve into the molecular mechanisms, e.g. SDF-1/CXCR4 and MMPs, which promote neuroblast migration in the ischemic brain after treatment with sildenafil. To examine the effects of sildenafil-induced angiogenesis on neurogenesis and neuroblast migration, we will measure the interaction between angiogenesis and neurogenesis and proteins secreted by the endothelial cells. By blocking SDF-1a and MMPs, we will examine whether SDF-1a and MMPs secreted by endothelial cells mediate neuroblast migration. Using specific inhibitors, we will investigate whether blocking the PI3K/Akt signaling pathway attenuates the effects of sildenafil on neurogenesis, neuroblast migration and angiogenesis. These basic scientific studies will elucidate the mechanisms how the brain remodels itself after stroke and how cGMP amplifies this process in the adult brain, which may lead to novel methods to facilitate brain remodeling during stroke recovery.