Developmental endothelial locus-1 (Del-1) is a hemostatic factor in thrombotic stroke

 描述(申请人提供)：AIS最常见的起因是血栓性血管闭塞，随后是一系列复杂的事件，包括炎症反应，共同导致缺血性神经元功能障碍。尽管进行了大量的研究，但组织型纤溶酶原激活剂(TPA)是唯一被批准的治疗AIS的方法。然而，短暂的治疗窗口和治疗后出血并发症限制了它的临床应用。因此，有必要在脑血栓形成和炎症的调节中寻找新的靶点，以将生理性止血和修复与病理性血栓形成分离开来。在这个建议中，我们关注最近发现的一种抗炎蛋白，发育内皮基因座-1(Del-1)，它在大脑中高表达，我们推测它也参与了导致AIS的血栓形成的发病机制。我们的数据表明，Del-1具有很强的抗血栓活性。我们认为，Del-1通过其磷脂酰丝氨酸(PS)结合域介导的抗血栓活性与被归因于整合素阻断的抗炎作用相结合。在缺血性血管损伤开始时，表达PS的细胞对Del-1的下调或摄取Del-1的增加而导致的一过性耗竭可能会加剧血栓的形成并增加梗塞面积。我们的初步数据首次显示，结构性Del-1缺乏症在卒中小鼠模型中是前血栓形成的，增加了梗塞面积，并导致更广泛的坏死。用我们在NM浓度下用人免疫球蛋白Fc片段构建的重组(R)-Del-1融合蛋白干预，可以挽救Del-/-小鼠的表型。我们的主要假设是Del-1是一种新的脑源性止血因子，在AIS的发病机制和潜在的治疗中发挥作用。这将在两个相互关联的目标中进行测试：目标1。我们将使用两种血管损伤模型、测量梗塞面积的MRI成像技术和皮质血流量来更全面地描述Del-1缺乏对血栓形成和AIS发展的影响。目的2.利用125I-纤维蛋白原和FITC-葡聚糖外渗，探讨r-Del-1-Fc在防止病理性血栓继发扩张的同时最小化止血作用的可能性。在后续的研究中，我们将阐明PS和整合素结合结构域在Del-1‘S在AIS发生中的作用。该项目的成功完成将确认Del-1是一种新的脑血管内皮细胞来源的动态平衡抗血栓蛋白，与AIS的发病机制有关。尽管这些研究是初步的，但Del-1对病理性血栓形成和炎症的潜在影响使其成为帮助防止中风扩大和复发的有吸引力的新候选药物。

 DESCRIPTION (provided by applicant): AIS is most commonly initiated by thrombotic vascular occlusion, followed by a complex series events including an inflammatory response that together leads to ischemic neuronal dysfunction. Notwithstanding intensive research, tissue-type plasminogen activator (tPA) is the only approved treatment for AIS. However, the brief therapeutic window and post-treatment bleeding complications have limited its clinical use. Therefore, there is a need to identify novel targets in the regulation of cerebral thrombosis and inflammation to dissociate physiological hemostasis and repair from pathological thrombus formation. In this proposal we focus on a recently discovered anti-inflammatory protein, developmental endothelial locus-1 (Del-1), which highly expressed in the brain that we hypothesize is also involved in the pathogenesis of thrombosis that leads to AIS. Our data demonstrate that Del- 1 has potent anti-thrombotic activity. We propose that Del-1 combines anti-thrombotic activity mediated through it phosphatidylserine (PS) binding domain with anti-inflammatory actions that have been attributed to integrin blockade. Down-regulation of Del-1 or transient depletion by increased uptake of Del-1 by PS-expressing cells at the onset of ischemic vascular injury may exacerbate thrombus formation and increase infarct size. Our preliminary data show for the first time that constitutive Del-1 deficiency is prothrombotic, increase infarct size and causes more extensive necrosis in a murine model of stroke. Intervention with a single dose of a fusion protein we created recombinant (r)-Del-1 with human IgG Fc fragment at nM concentrations rescues the phenotype of Del-/- mice. Our main hypothesis is that Del-1 is a novel brain derived hemostatic factor that plays a role in the pathogenesis and potentially the management of AIS. This will be tested in two interrelated aims: Aim 1. We will more fully characterize the impact of Del-1 deficiency on the development of thrombosis and AIS using two models of vascular injury, MRI imaging techniques to measure infarct size, and cortical blood flow. Aim 2. We will explore the potential of r-Del-1-Fc to prevent secondary expansion of pathological thrombosis while minimizing effects on hemostasis using 125I-fibrinogen and FITC-dextran extravasation. In follow-on studies, we will delineate the contribution of the PS and integrin binding domains to Del-1's role in the development of AIS. Successful completion of this project will identify Del-1 as a novel cerebrovascular endothelium derived homeostatic anti-thrombotic protein relevant to the pathogenesis of AIS. Although these studies are preliminary, the potential impact of Del-1 on pathological thrombosis and inflammation makes it an attractive new candidate to help prevent stroke expansion and recurrence.