Cell-Cell Interactions in Thrombosis -Request for Extension of Merit Award

冠状动脉、脑动脉和外周动脉损伤可引起局部血小板活化、重新聚集和血栓形成 遮挡。预防和治疗血小板驱动的血栓形成是一项治疗挑战，关键是 对公共健康的影响。在内皮细胞存在的情况下，血小板对激动剂没有反应。这个没有回应的- 内皮型NTPDasel/CD39/ecto-ADPase能迅速代谢ATP，释放ADP 来自激活的血小板。这取消了聚合和招聘。重组人可溶性CD39(SolCD39) 在v#ro中阻断人的血小板聚集，并在体外抑制猪和常规的血小板聚集。CD39为空 小鼠表现出潜在的血栓前表型，对中风、心脏和肺的易感性增加 血栓形成。通过注射solCD39，野生型和CD39缺失小鼠的这种情况都得到了缓解，显示了其关键的 在血栓调节中的作用。广泛的长期目标和具体目标包括：i分子、生化和 人solCD39的功能研究：经筛选的突变体抗血栓效果的测定 ADP的特异性/活性，以及二硫键对CD39活性的贡献的评估。11影响的研究 缺血时NTPDase对细胞-细胞信号的影响：心脏NTPDase对去甲肾上腺素释放的调节 豚鼠、野生型和CD39缺失小鼠的交感神经末梢(CSNE)和离体心。鉴定 小鼠和人心脏cSNE胞外核苷酸酶补体的测定及其肺保护作用的评价 通过内源性和外源性CD39在第二器官灌流损伤过程中，野生型和CD39缺失小鼠。I]1 冠状动脉造影确诊患者淋巴细胞中NTPDase活性的研究 疾病(CAD)，以确定是否表达改变的，血栓前核苷酸酶与 来自健康捐献者的淋巴细胞。CD39产生的AMP腺苷的E代谢 ATP和ADP：PRP可通过产生腺苷代谢AMP并抑制血小板活性 5‘-核苷酸酶？如果是真的，这是依赖于特定的血小板激动剂，还是依赖于其他血液或内皮细胞？二恶英 毒性可能包括通过上调腺苷脱氨酶增加腺苷的分解代谢，从而减少 一种有效的血小板抑制剂腺苷的可用性。这项研究代表了一种多学科的方法来 了解CD39作为血小板介导的闭塞性血栓形成的主要调节因子的关键作用。它是基于 关于令人信服的可行性数据和历史合作成功，并将促进对NTPDase的理解 生物学和血栓调节，最终形成了一种治疗血栓性疾病的独特和新颖的治疗剂。

Injury to coronary, cerebral, and peripheral arteries evokes local platelet activation, recruitment, and thrombotic occlusion. Prevention and treatment of platelet-driven thrombus formation is a therapeutic challenge, with critical public health implications. In the presence of endothelial cells, platelets are unresponsive to agonists. This unrespon- siveness is due to endothelial NTPDasel/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 v#ro, and inhibits porcine and routine 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. Broad, long-term objectives and specific aims include: I Molecular, biochemical and functional studies of human solCD39: Determination of the antithrombotic efficacy of selected mutants with increased ADP specificity/activity, and evaluation of the contribution of disulfide bridges to CD39 activity. 11Studies of the effects of NTPDases on cell-cell signaling during ischemia: Modulation of norepinephrine release by NTPDases from cardiac sympathetic nerve endings (cSNE) and isolated hearts from guinea pigs, wild type and CD39 null mice. Identification of the ecto-nucleotidase complement in cSNE from mouse and human heart, and assessment of pneumoprotection by endogenous and exogenous CD39 during second organ perfusion injury, inwild-type and CD39 null mice. I]1 Studies of NTPDase activities in lymphocytes from patients with angiographically documented coronary artery disease (CAD) to determine whether an altered, prothrombotic nucleotidase profile is expressed in comparison to that of lymphocytes from healthy donors. E Metabolism of adenosine derived from AMP as generated by CD39 from ATP and ADP: Can PRP metabolize AMP and generate inhibition of platelet reactivity by production of adenosine via 5'-nucleotidase? If true, is this dependent on specific platelet agonists, or on other blood or endothelial cells? Dioxin toxicity may involve increased catabolism of adenosine via upregulation of adenosine deaminase, thus reducing availability of adenosine, a potent platelet inhibitor. The research represents a multidisciplinary approach to understanding the critical role of CD39 as the prime regulator of platelet-mediated occlusive thrombosis. It is based on compelling feasibility data and historical collaborative success, and will advance the understanding of NTPDase biology and thromboregulation, culminating in a unique and novel therapeutic agent for thrombotic diatheses.