Ectonucleotidases in Atherothrombosis and Stroke

描述(申请人提供)：激活的血小板释放的ADP招募附近的血小板，导致血栓性病灶的爆炸性增加。激活的血小板或受损细胞释放的三磷酸腺苷促进炎症。胞外核苷酸在胞外核苷酸酶CD39(核苷酸二磷酸水解酶1)的顺序作用下，协同磷酸水解酶将ATP？ADP？AMP)和CD73(51核苷酸酶，转化为AMP？腺苷)，是一种重要的内皮稳态机制，限制血管界面的血栓形成和炎症。CD39基因缺失的小鼠在局灶性脑缺血的背景下表现出潜在的血栓前表型和比对照组更差的结果。这些小鼠可以通过保留apyrase活性的重组可溶性CD39来拯救，而不会增加脑内出血。此外，高胆固醇血症ApoE/CD39双基因敲除小鼠表现出过度的动脉粥样硬化，这与血小板和炎症细胞重新聚集到发展中的斑块中的作用一致。这些数据表明，胞外核苷酸酶可预防动脉粥样硬化血栓形成事件，并与中风的发病机制有关。实验将阐明CD39和CD73具有动脉粥样硬化保护作用的机制，重点是在对照或高胆固醇血症背景下，使用野生型CD39或CD73基因缺失的小鼠，研究它们抑制血小板激活和炎症级联的能力。实验将使用solCD39(和/或纯化的CD73)作为单一疗法或作为低剂量溶栓疗法的辅助手段，测试ectDNA酶是否能在容易发生动脉粥样硬化的脑血管环境中改善缺血性中风的预后。这里的首要目标是描绘保护血管免受动脉粥样硬化血栓形成的内源性级联反应，并确定它是否可以在治疗上用于改善缺血性卒中的结果。 与公共卫生相关：该项目将探索存在于血管衬里细胞上的两种蛋白质如何降解循环物质，否则会促进凝血、炎症和动脉粥样硬化。利用这些蛋白质的活性可能会导致一种防止血栓形成和动脉粥样硬化的新方法，从而导致一种全新的、或许更安全的中风治疗方法。

DESCRIPTION (provided by applicant): ADP released from activated platelets recruits nearby platelets, resulting in explosive accretion of a thrombotic nidus. ATP, released from activated platelets or injured cells, promotes inflammation. The coordinated phosphohydrolysis of extracellular nucleotides, by the sequential actions of the ectonucleotidases CD39 (nucleotide diphosphohydrolase 1, converts ATP? ADP?AMP) and CD73 (51 nucleotidase, converts AMP?adenosine), is an important endothelial homeostatic mechanism which limits thrombosis and inflammation at the blood-vessel interface. CD39 gene null mice exhibit a latent prothrombotic phenotype and worse outcomes than controls in the setting of focal cerebral ischemia. These mice can be rescued by recombinant soluble CD39, which retains apyrase activity, without increasing intracerebral hemorrhage. Furthermore, hypercholesterolemic ApoE/CD39 double knockout mice exhibit exaggerated atherogenesis, consistent with a role for platelet and inflammatory cell recruitment into the developing plaque. These data suggest that ectonucleotidases protect against atherothrombotic events and are relevant to the pathogenesis of stroke. Experiments will elucidate mechanisms by which CD39 and CD73 are atheroprotective, focusing on their ability to suppress platelet activation and inflammatory cascades, using wild-type, cd39- or cd73-gene null mice in control or hypercholesterolemic backgrounds. Experiments will test whether ectonucleotidases improve ischemic stroke outcomes in an atherosclerosis-prone cerebrovascular milieu, using solCD39 (and/or purified CD73) as monotherapy or as an adjunct to low dose thrombolytic therapy. The overarching goal here is to delineate an endogenous cascade which protects vessels against atherothrombosis, and determine whether it may be therapeutically harnessed to ameliorate ischemic stroke outcomes. Relevance to Public Health: This project will explore how two proteins, which exist on cells lining blood vessels, degrade circulating substances which would otherwise promote clotting, inflammation, and atherosclerosis. Harnessing the activity of these proteins might lead to a new way of preventing clot formation and atherosclerosis, and thereby lead to a completely new and perhaps safer treatment for stroke.