Biomedical and Functional Studies of CD39

冠状动脉、脑动脉或外周动脉损伤引起局部血小板活化、募集， 血栓性闭塞预防和治疗血小板驱动的血栓形成是一种治疗方法 这是一项重大挑战，对公共卫生有重大影响。在存在内皮细胞的情况下，血小板被 对激动剂无反应这种无反应性是由于内皮NTPD酶1/CD 39/外膜-ADVANE， 其快速代谢从活化的血小板释放的ATP和ADP。这废除了聚合， 招聘重组可溶性人CD 39（solCD 39）在体外阻断人血小板聚集， 抑制猪和鼠的离体血小板聚集。CD 39敲除小鼠表现出潜在的血栓前病变， 表型与增加的中风和心脏和肺血栓形成的易感性。这是缓解 在野生型和CD 39缺失小鼠中，通过输注solCD 39，证实了其在 血栓调节该项目的长期目标和具体目标包括：一代大 用于生化和功能研究的重组可溶性CD 39的数量，包括 血管损伤和闭塞，使用我们的CD 39敲除小鼠。这些型号包括：a）经腔导丝 损伤小鼠股动脉，其中我们将确定CD 39在初始血小板- 依赖性阶段以及该血管损伤的晚期增殖阶段;和B}鼠LAD结扎 导致心肌缺血/再灌注损伤。这将描述solCD 39在治疗中的有效性。 短期心肌缺血/再灌注（与solCD 39在减少心肌缺血/再灌注损伤的大小方面的有效性相当）。 CD 39基因敲除小鼠脑梗死的发生率），以及长期条件（LV重塑和充血性心脏病 失败）。[L]心脏和外周中嘌呤能和肾上腺素能神经传递的调节 NTPDase 1/CD 39在缺血时对交感神经系统的影响。这些研究将阐明 CD 39在心脏交感神经传递中的作用，并证明了CD 39在心脏交感神经传递中的重要调节作用。 外周交感神经效应器接头。此外，他们将验证ATP和NE的溢出， 再灌注心律失常的严重程度可以被减弱。因此，由CD 39提供的心脏保护作用可以是 在CD 39缺失动物中恢复。IN.使用CD和NMR的可溶性腺苷三磷酸双磷酸酶的结构分析将建立 蛋白质-核苷酸相互作用，结合动力学，并描绘了活性位点的重要方面， solCD 39-主要的血栓调节剂。这项研究是一项多学科的研究， 合作的方法来理解CD 39作为血液流动性的主要调节剂的关键作用， 血小板介导的闭塞性血栓形成这些研究基于令人信服的可行性数据， 历史性的合作成功，并将促进对NTPDase生物学的理解， 血栓调节这将最终导致一种独特的和新的治疗血栓性疾病的药物。

Injury to coronary, cerebral, or peripheral arteries evokes local platelet activation, recruitment, and thrombotic occlusion. Prevention and treatment of platelet-driven thrombus formation is a therapeutic challenge, with major public health implications. In the presence of endothelial cells, platelets are unresponsive to agonists. This unresponsiveness is due to endothelial NTPDase1/CD39/ecto-ADPase, which rapidly metabolizes ATP and ADP released from activated platelets. This abolishes aggregation and recruitment. Recombinant, soluble human CD39 (solCD39) blocks human platelet aggregation in vitro, and inhibits porcine and murine platelet aggregation ex vivo. CD39 null mice exhibit a latent prothrombotic phenotype with increased susceptibility to stroke and cardiac and pulmonary thrombosis. This is alleviated in both wild-type and CD39 null mice by infusion of solCD39, demonstrating its critical role in thromboregulation. Long-term objectives and specific aims of this project include: I. Generation of large quantities of recombinant soluble CD39 for biochemical and functional studies, including mouse models of vascular injury and occlusion, employing our CD39 null mice. These models include: a) Transluminal wire injury of mouse femoral artery, in which we will ascertain the role of CD39 in both the initial platelet- dependent stage, as well as the late proliferative stage of this vascular injury; and b} murine LAD ligation leading to myocardial ischemia/reperfusion injury. This will delineate the effectiveness of solCD39 in myocardial ischemia/reperfusion in short-term (comparable to effectiveness of solCD39 in reducing the size of brain infarct in CD39 null mice), as well as long-term conditions (LV remodeling and congestive heart failure). [L Modulation of purinergic and adrenergic neurotransmission in the heart and peripheral sympathetic nervous system by NTPDase1/CD39 during ischemia. These studies will elucidate the role of CD39 in sympathetic transmission in the heart and demonstrate the crucial modulatory role of CD39 at peripheral sympathetic neuro-effector junctions. Moreover, they will verify that overflow of ATP and NE, and severity of reperfusion arrhythmias, can be attenuated. Thus, cardioprotection afforded by CD39 can be restored in CD39 null animals. IN. Structural analyses of soluble apyrases using CD and NMR will establish protein-nucleotide interactions, kinetics of binding, and delineate important aspects of the active site of solCD39 - the principal thromboregulator. The proposed research represents a multidisciplinary and collaborative approach to understanding the critical role of CD39 as the prime regulator of blood fluidity and of platelet-mediated occlusive thrombosis. The studies are based on compelling feasibility data and historical collaborative success, and will advance the understanding of NTPDase biology and thromborequlation. This should culminate in a unique and novel therapeutic aqent for thrombotic diatheses.