DIFFERENCES IN THE PROTEIN SIGNATURES/PNH PLATELETS

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 阵发性睡眠性血红蛋白尿症是一种获得性溶血性贫血， 通过增加红细胞对补体的敏感性， 溶血和血红蛋白尿。PNH是由于细胞克隆的扩增， 在X连锁的PIGA基因中获得了突变。PIGA是一种必需的酶亚基， 用于合成糖基磷脂酰肌醇（GPI）锚分子。血细胞 因此，来自突变祖细胞的所有GPI锚定的细胞都是缺陷的。 分子。我们研究的广泛长期目标是了解 PNH的病理生理和发病机制。PNH是一种慢性疾病， 发病率和死亡率都很高血栓形成是最常见的原因， 死亡PNH中血栓形成的病理生理机制尚不清楚。我们提出 GPI连接蛋白（PNH表型）缺陷的血小板（Plt）在PNH中起主要作用。 在PNH血栓形成的发病机制中的作用。我们假设， PNH表型不仅缺乏所有GPI-连接蛋白， 其他蛋白质，其合成或定位依赖于正常GPI锚 生产，这些蛋白质的缺陷可能有助于 血栓前风险。在拟议的研究中，我们将重点关注分子 通过鉴定PNH中的蛋白质和蛋白质修饰， 与PNH相关的血小板。首先，我们将开发可重复的蛋白质组学 从正常和PNH患者中分离和分析Plt的程序，然后 将来自GPI连接蛋白缺陷的Plts的蛋白质谱与来自GPI连接蛋白缺陷的Plts的蛋白质谱进行比较， 正常皮特的侧写最后，我们将开发检测和测量 PNH细胞特异性的候选蛋白或蛋白修饰，并验证 第二组患者和对照中候选蛋白的差异表达 个体我们建议的调查可能会发现一些 意想不到的蛋白质，蛋白质相互作用和蛋白质修饰， 显著提高了我们对PNH血栓形成发病机制的认识 并且也可能在其它病症中，其中异常克隆造血被 与血栓形成相关，例如真性红细胞增多症或其他 骨髓增生综合征建立了分析和比较 血小板蛋白质组在健康和疾病可能导致识别新的 用于评估血栓形成风险或鉴定新靶点的生物标志物 诊断或药物的发展。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Paroxysmal nocturnal hemoglobinuria is an acquired hemolytic anemia characterized by the increased sensitivity of red cells to complement leading to intravascular hemolysis and hemoglobinuria. PNH is due to the expansion of a cell clone that has acquired a mutation in the X-linked PIGA gene. PIGA is an enzyme subunit essential for the synthesis of glycosyl phosphatidylinositol (GPI) anchor molecules. Blood cells derived from the mutant progenitor cell are therefore deficient in all GPI-anchored molecules. The broad long-term objective of our research is to understand the pathophysiology and pathogenesis of PNH. PNH is a chronic disease often associated with substantial morbidity and mortality. Thrombosis is the most frequent cause of death. The pathophysiology of thrombosis in PNH is not understood. We propose that platelets (Plt's) deficient in GPI-linked proteins (PNH phenotype) play a major role in the pathogenesis of thrombosis in PNH. We hypothesize that blood cells with the PNH phenotype not only lack all GPI-linked proteins, but are also deficient in other proteins, whose synthesis or localization is dependent on normal GPI anchor production, and that the deficiencies of these proteins on Plt's might contribute to the prothrombotic risk. In the proposed research we will focus on the molecular aspects of these hypotheses by identifying proteins and protein modification in PNH Plt's that are associated with PNH. First, we will develop reproducible proteomic procedures to isolate and analyze Plt's from normal and PNH patients, and then compare the protein profile from Plts' deficient in GPI-linked proteins with the protein profile of normal Pit's. Finally, we will develop assays to detect and measure candidate proteins or protein modifications specific for PNH cells and verify the differential expression of candidate proteins in a second cohort of patient and control individuals. Our proposed investigations are likely to identify a number of unanticipated proteins, protein interactions, and protein modifications that might significantly advance our understanding of the pathogenesis of thrombosis in PNH and possibly also in other conditions, in which an abnormal clonal hematopoiesis is associated with thrombosis, for example, polycythemia vera or other myeloproliferative syndromes. Established methods for analyzing and comparing the platelet proteome in health and disease might lead to the identification of novel biomarkers useful in evaluating the risk of thrombosis or identify new targets for the development of diagnostics or drugs.