The Impact of Hemoglobin S on Red Blood Cell Nitric Oxide Production

项目摘要 镰状细胞病是一种进行性血管疾病，源于红细胞变形性降低。 血管疾病是急性和慢性镰状疾病的核心，包括疼痛危象、急性胸部 综合症、中风、皮肤溃疡和肺动脉高压。然而，连接机制的数量减少了 红细胞对慢性血管病变的变形能力是多因素的，且特征不佳。一氧化氮(NO)是 将血液力学与血管张力和血管重塑联系起来的关键介质。生物利用度不会降低 在SCD中，由于溶血过程中释放的脱细胞血红蛋白和精氨酸酶，清除NO和更低的 血管内皮细胞产生NO。最近的证据表明，50%的生物可用NO是在红细胞内合成的， 通过剪切激活的内皮型一氧化氮合酶。RBC NO主要转化为亚硝酸盐 当组织含氧量高时，硝化血红蛋白，但脱氧血红蛋白将这些转化为 在低氧条件下，物种对一氧化氮的反应。因此，RBC产生的NO似乎是一种重要的调节因子。 氧的供需及其在镰状细胞血管病变中的作用尚不清楚。我们实验室的初步结果 提示组织氧合依赖于红细胞在高切变力下的变形能力。健康和变形的人 SCD RBC对NO产生的促进作用相似，而SCD RBC基础NO的产生高于SCD RBC。使用 在完全氧化的SCD红细胞中加入亚硝酸盐增加了基础NO的产生，而不是 健康红细胞的变化。我们的总体目标是证明亚硝酸盐和一氧化氮合酶对红细胞一氧化氮的贡献 产生，进而对正常健康受试者的血管健康起着重要作用 镰状细胞病患者，一种人类弥漫性血管病变的模型。这项研究建议 利用我们目前在镰状细胞疾病血管功能评估方面的工作和新的实验室方法 红细胞产生一氧化氮。不同血管床的多模式表征将导致改进 表型分类和与潜在红细胞生物物理/生化的病理生理联系 精神错乱。我们继续探索RBC产生的NO是否具有降低血小板的能力 聚合。目标一和目标二中提出的研究将分离基础和剪切介导的一氧化氮的影响。 生产使我们能够确定健康和SCD患者的控制机制。我们知道，一个 在未输血的SCD受试者中，存在组织氧合低，而微循环血流的悖论 是增加的。这可能是由于S血红蛋白还原亚硝酸盐导致一氧化氮产生的变化。 脱氧，剪切力介导的NO产生的变化或两者兼而有之。我们的整体设计，实现了目标 I/II与目标III的研究同时进行，应能解决这一矛盾。K23机制代表着 自然地扩展了我迄今为止的职业发展，结合了我以前的实验室和以病人为中心 具有进行大型转化性研究所需的特定临床研究培训的研究专业知识 血管功能障碍的新靶点。

Project Summary Sickle cell disease is a progressive vasculopathy stemming from decreased red blood cell (RBC) deformability. Vascular disease is at the heart of both acute and chronic sickle disease, including pain crisis, acute chest syndrome, stroke, skin ulcers, and pulmonary hypertension. However, the mechanisms linking decreased RBC deformability to chronic vasculopathy are multifactorial and poorly characterized. Nitric oxide (NO) is the key mediator linking blood mechanics to vessel tone and vascular remodeling. NO bioavailability is diminished in SCD because decellularized hemoglobin and arginase, released during hemolysis, scavenge NO and lower endothelial NO production. Recent evidence suggests that 50% of bioavailable NO is synthesized within RBC, themselves, through a shear-activated eNOS enzyme. RBC NO is primarily converted to nitrite and nitrosylated hemoglobins when tissue oxygenation is high, but deoxygenated hemoglobin converts these species to nitric oxide under hypoxic conditions. Thus, RBC generated NO appears to be a vital mediator of oxygen supply and demand and its role in sickle cell vasculopathy is unexplored. Early results from our lab suggest that tissue oxygenation is dependent on RBC deformability at high shear. Deformation of healthy and SCD RBC increases NO production to a similar degree, while basal NO production is higher in SCD RBC. With the addition of nitrite to fully oxygenated SCD RBC basal production of NO is increased whereas it did not change in healthy RBC. Our overall goal is to demonstrate that nitrite and NOS contribute to RBC NO production, which in turn plays a significant role in the vascular health of normal healthy subjects and patients with sickle cell disease, a human model of diffuse vasculopathy. This research proposal leverages our current work in sickle cell disease vascular function assessment and novel laboratory methods in RBC nitric oxide production. Multimodal characterization of the different vascular beds will lead to improved phenotypic categorization and pathophysiological links to the underlying RBC biophysical/biochemical derangements. We continue to explore whether RBC-generated NO has the ability to decreases platelet aggregation. The studies proposed in Aim I and II will separate the effect of basal and shear-mediated NO production allowing us to determine control mechanisms in healthy and SCD patients. We know that a paradox exists whereby tissue oxygenation is low in non-transfused SCD subjects, while microcirculatory flow is increased. This may be due to changes in nitric oxide production due to nitrite reduction from hemoglobin S deoxygenation, shear-mediated changes in NO production or both. Our overall design, which performs Aims I/II simultaneously with studies in Aim III, should resolve this paradox. The K23 mechanism represents the natural extension my career development to date, combining my previous laboratory and patient-oriented research expertise with the specific clinical research training necessary to conduct large translational studies of novel targets in vascular dysfunction.