Regulation of Stromal Derived Factor-1 Mediated Angiogenesis in Ischemic Brain

描述（由申请人提供）：中风是发病和长期残疾的第三大原因。由于脑动脉闭塞，缺血性中风导致血液供应严重减少，导致内皮完整性破坏和神经元细胞死亡。事实上，脑血管密度较高的患者在中风后表现出更好的恢复和生存。许多炎性趋化因子支持血管血液供应（血管生成）的发展和祖细胞向损伤部位的迁移。特别是，趋化因子基质衍生因子（SDF 1）通过其受体CXCR 4发挥作用，在促进血管生成和祖细胞募集中发挥核心作用。然而，SDF 1通常被蛋白水解切割和失活。这一过程可能会阻碍神经（NPC）和内皮祖细胞（EPC）的迁移和脑损伤修复所需的血管生成。因此，防止SDF 1失活具有临床重要性。虽然蛋白酶二肽基肽酶4（DPPIV）被证明可以切割SDF 1，但其在缺血性卒中中的作用尚不清楚。该提案的目标是建立一个基础工作，用于评估DPPIV抑制在增强SDF 1活性以改善血管生成和脑损伤修复方面的功效。我们的研究表明DPPIV的缺失和SDF 1水平的增加之间存在相关性，导致神经嵴干细胞衍生的神经母细胞瘤细胞的迁移和血管生成潜力增加。我们进一步观察到缺血后脑中DPPIV表达显著增加。我们推测，局灶性缺血后，DPPIV上调减少SDF 1活性，从而阻碍祖细胞迁移到缺血区域，并抑制随后的血管生成和神经发生。我们进一步提出DPPIV的基因敲除或小分子抑制剂增加缺血后SDF 1水平，这反过来又增强了缺血脑中NPC和EPC的迁移和血管生成。在本提案中，我们将测试短暂性大脑中动脉闭塞（MCAO）后小鼠脑和血清中DPPIV表达与SDF 1和CXCR 4水平之间的预测负相关性（目的1）。我们将研究是否DPPIV或DPPIV的小分子抑制剂的遗传损失增强EPC招聘和血管生成在体内缺血性脑和体外缺血模型的氧葡萄糖剥夺。将表达荧光素酶的NPC/EPC移植到经受MCAO的小鼠的对侧纹状体中，并使用生物发光成像追踪迁移细胞（Aim 2）。重要的是，DPPIV抑制剂是FDA批准的抗糖尿病药物，可增加糖尿病患者的循环EPCs。我们的研究如果成功，可以转化为临床前和临床研究，以改善血管生成和神经发生。这些研究结果可能为更好地理解脑损伤修复的新靶点和机制提供强有力的基础，也可能为中风治疗开辟新的方向。 公共卫生相关性：缺血性卒中导致死亡或长期残疾，对患者和护理人员提出了独特的挑战。中风导致血流量突然减少和神经细胞死亡。恢复血流和神经发生对中风恢复至关重要。目前，治疗中风患者的治疗选择有限。我们的研究代表了更好地理解缺血后大脑中控制神经发生和血管生成（新血管发芽）的新分子事件的重要第一步，并为进一步测试DPPIV小分子抑制剂的功效奠定了基础缺血性中风后改善脑损伤修复仍然是世界上的主要健康问题。

DESCRIPTION (provided by applicant): Stroke is the third leading cause of morbidity and long-term disability. Due to cerebral artery occlusion, ischemic stroke causes a severe reduction in blood supply causing destruction of endothelial integrity and neuronal cell death. Indeed, patients with higher density of cerebral blood vessel show better recovery and survival after stroke. Many inflammatory chemokines support the development of vascular blood supply (angiogenesis) and progenitor cell migration to the site of injury. Particularly, the chemokine stromal derived factor (SDF1) acting via its receptor CXCR4 plays a central role in promoting angiogenesis and progenitor cell recruitment. However, SDF1 is often proteolytically cleaved and inactivated. This process may hinder neural (NPC) and endothelial progenitor cells (EPC) migration and angiogenesis necessary for brain injury repair. Thus preventing SDF1 inactivation is of clinical importance. Although the protease dipeptidyl peptidase 4 (DPPIV) is shown to cleave SDF1, its role in ischemic stroke is unknown. The goal of this proposal is to establish a ground work for evaluating the efficacy of the DPPIV inhibition in enhancing the activity of SDF1 for improved angiogenesis and brain injury repair. Our studies show a correlation between loss of DPPIV and increased levels of SDF1 resulting in increased migratory and angiogenic potential of neural crest stem cell derived neuroblastoma cells. We further observed significantly increased DPPIV expression in the post- ischemic brain. We hypothesize that following focal ischemia, DPPIV up regulation curtails SDF1 activity and thus hinders the migration of progenitor cells to the ischemic region and suppresses subsequent angiogenesis and neurogenesis. We further propose that genetic knockout or small molecule inhibitors of DPPIV increases post-ischemic SDF1 levels, which in turn enhances NPC and EPC migration and angiogenesis in ischemic brain. In this proposal, we will test the predicted inverse correlation between DPPIV expression and SDF1, and CXCR4 levels in mouse brain and serum following transient middle cerebral artery occlusion (MCAO) (Aim 1). We will examine whether genetic loss of DPPIV or small molecule inhibitor of DPPIV enhances EPC recruitment and angiogenesis in vivo ischemic brain and in an in vitro ischemic model of oxygen glucose deprivation. The luciferase expressing NPC/EPC will be transplanted into the contralateral striatum of the mice subjected to MCAO and migrating cells will be tracked using bioluminescence imaging (Aim 2). Importantly, DPPIV inhibitors are FDA approved anti-diabetic drugs that increase circulating EPCs in diabetic patients. Our studies if successful can be translated to pre-clinical and clinical studies for improved angiogenesis and neurogenesis. Results from these studies may provide strong foundation for better understanding of novel targets and mechanisms of brain injury repair and may also open up a new direction for stroke therapy. PUBLIC HEALTH RELEVANCE: Ischemic stroke causes death or long term disability presenting a unique challenge to patients and care givers. Stroke results in sudden decrease in blood flow and neural cell death. Restoring blood flow and neurogenesis is critical for stroke recovery. At present, there are limited therapeutic options to treat stroke patients. Our study represents an important first step in better understanding the novel molecular events that govern neurogenesis and angiogenesis (sprouting of new blood vessels) in the post-ischemic brain and establishes ground work for further testing the efficacy of small molecule inhibitors of DPPIV for improved brain injury repair after ischemic stroke that remains a major health problem in the world.