Mechanisms of platelet stimulation in hemolysis

溶血中血小板刺激的机制

• 批准号: 10571830
• 负责人: Deirdre Nolfi-Donegan
• 金额: $ 12.82万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-15 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10571830
• 关键词: Advisory Committees; Attenuated; Blood; Blood Platelet Disorders; Blood Platelets; Blood Vessels; Cells; Complex; Cysteine; Data; Development; Development Plans; Disease; Disulfides; Electron Transport; Environment; Erythrocytes; Event; Experimental Models; Foundations; Generations; Genetic Diseases; HMGB1 gene; Haptoglobins; Heme; Hemoglobin; Hemolysis; Hemopexin; Human; Hypoxia; Incubated; Inflammation; Inflammation Mediators; Inflammatory; Infusion procedures; Institute of Medicine (U.S.); Knock-out; Life; Link; Mass Spectrum Analysis; Measurement; Measures; Mediating; Membrane Potentials; Mentors; Mentorship; Mitochondria; Molecular; Mus; Oxidation-Reduction; Pathogenesis; Pathway interactions; Patients; Pattern; Plasma; Platelet Activation; Platelet aggregation; Play; Polymers; Production; Proteins; Pulmonary Hypertension; Reactive Oxygen Species; Receptor Signaling; Recombinants; Research; Research Personnel; Respiration; Risk; Role; Sickle Cell Anemia; Sickle Hemoglobin; Signal Pathway; Signal Transduction; Sterility; Stimulant; Stimulus; Stroke; Sulfhydryl Compounds; Surface; TLR4 gene; Therapeutic; Thrombosis; Thrombus; Training; Transgenic Mice; Universities; Work; career; career development; ferric chloride; hemoglobin polymer; high risk; improved; in vivo; inhibitor; mouse model; multidisciplinary; mutant; new therapeutic target; novel; oxidation; prevent; receptor; synergism; therapeutic target; thrombotic; thrombotic complications

ABSTRACT While sickle cell disease (SCD) is instigated by a mutant hemoglobin that polymerizes in hypoxia to cause hemolysis and vaso-occlusion, inflammation plays a fundamental role in propagating this pathogenesis. SCD patients are at significantly higher risk of thrombosis, which contributes to severe life-threatening complications such as pulmonary hypertension and stroke. On a cellular level, it is well established that in SCD patients, platelet activation is elevated and correlates with hemolysis. However, the contribution of inflammatory signaling to platelet activation and its cross-talk with hemolysis remain unknown. High-mobility group box 1 (HMGB1), an inflammatory mediator released by cells, has been shown to be elevated in the blood of SCD patients and to stimulate platelet activation. Notably, the ability of HMGB1 to mediate platelet activation is dependent on the oxidation state of critical cysteine residues in the protein. Preliminary data shows that HMGB1-mediated platelet activation is significantly enhanced in the presence of cell-free hemoglobin (Hb), a major component of hemolysis. Further, Hb and HMGB1 each independently increase platelet mitochondrial reactive oxygen species (mtROS) generation, a known stimulus for platelet activation. Based on these data, I hypothesize that in SCD, Hb oxidizes HMGB1 to increase platelet mtROS production leading to platelet activation and secondary release of additional thrombotic stimulants including HMGB1 from platelets. Aim 1 will determine whether Hb oxidizes HMGB1 to potentiate its activation of platelets using human healthy and sickle Hb and platelets in ex vivo experimental models. Aim 2 will determine the mechanism by which HMGB1-induced mtROS stimulates platelet activation. Aim 3 will utilize murine models of hemolysis to determine whether neutralizing HMGB1 attenuates platelet activation and thrombus formation. Successful completion of these aims will elucidate a novel mechanism of synergy between hemolysis and inflammatory signaling in potentiating thrombosis. This work will reveal novel therapeutic targets and potential therapeutics for the treatment of thrombosis in SCD. In addition, this project, along with the guidance of my multi-disciplinary mentorship team, career development plan, and cutting-edge research environment at the Vascular Medicine Institute at the University of Pittsburgh, will serve as a strong training vehicle to facilitate my independence as a researcher focused on mechanisms of thrombosis in hemolytic disease.

摘要 虽然镰状细胞病（SCD）是由一种突变的血红蛋白引起的，这种血红蛋白在缺氧时聚合， 溶血和血管闭塞，炎症在传播这种发病机制中起着重要作用。SCD 患者发生血栓形成的风险明显更高，这会导致严重危及生命的并发症 例如肺动脉高压和中风。在细胞水平上，已充分确定在SCD患者中，血小板减少， 活化升高并与溶血相关。然而，炎症信号的贡献， 血小板活化及其与溶血的相互作用仍然未知。高迁移率族蛋白1（HMGB1）， 由细胞释放的炎性介质，已经显示在SCD患者的血液中升高， 刺激血小板活化。值得注意的是，HMGB1介导血小板活化的能力依赖于血小板活化。 蛋白质中关键半胱氨酸残基的氧化态。初步数据显示，HMGB1介导的血小板 在无细胞血红蛋白（Hb）的存在下，活化显著增强， 溶血此外，Hb和HMGB1各自独立地增加血小板线粒体活性氧 （mtROS）生成，一种已知的血小板活化刺激。基于这些数据，我假设在SCD中， Hb氧化HMGB1增加血小板mtROS产生，导致血小板活化和二次释放 额外的血栓形成刺激物，包括来自血小板的HMGB1。目标1将确定Hb是否氧化 使用人健康和镰状血红蛋白和血小板在离体中增强HMGB1对血小板的活化 实验模型目的2探讨HMGB1诱导线粒体活性氧刺激血小板的机制 activation.目的3将利用溶血的鼠模型来确定中和HMGB1是否减弱 血小板活化和血栓形成。成功地完成这些目标将阐明一部小说 溶血和炎症信号在增强血栓形成中的协同作用机制。这项工作将 揭示了新的治疗靶点和治疗SCD血栓形成的潜在疗法。此外，本发明还提供了一种方法， 这个项目，沿着我的多学科导师团队的指导，职业发展计划， 匹兹堡大学血管医学研究所的尖端研究环境将为 作为一个强有力的培训工具，以促进我作为一个专注于血栓形成机制的研究人员的独立性 溶血性疾病