IN VIVO ARGININE METABOLISM IN IDIOPATHIC AND SECONDARY PULMONARY HYPERTENSION

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。 摘要 肺动脉高压（PAH）是一种病因不明的疾病，其特征是肺动脉压力和血管阻力增加。它可以是特发性的或继发于其他疾病，如镰状细胞病。血管舒张分子一氧化氮（NO）的产生减少被认为在PAH的发病机制中起重要作用，但需要进一步在PAH患者中进行NO代谢的体内研究。我们认为，PAH患者中NO缺乏是由于NO前体精氨酸的可用性降低和一氧化氮合酶（合成NO的酶）活性受损的综合作用所致。具体而言，我们假设与健康对照组相比，PAH患者继发于精氨酸酶活性增加的可用于NO合成的精氨酸减少，将精氨酸分解为尿素和鸟氨酸的酶，并减少精氨酸的从头合成。此外，它们将具有增加的不对称二甲基精氨酸浓度，导致NO合酶活性的抑制。为了检验这些假设，将使用稳定同位素示踪技术测量健康对照、特发性PAH患者和继发于镰状细胞病的PAH患者的精氨酸和一氧化氮代谢。这些测量值将与血流动力学和临床参数相关。该项目将增加我们对肺动脉高压的发病机制中的作用，精氨酸-NO途径的知识。此外，它还可以确定未来治疗试验的目标。 I.假设 在这个项目中，我们建议测试的一般假设，减少血管舒张分子一氧化氮（NO）的可用性是肺动脉高压（PAH）的发病机制。这种NO的缺乏是由于NO前体精氨酸的可用性降低和一氧化氮合酶活性受损的综合作用导致的合成减少的结果。待检验的具体假设如下： 1.与健康对照相比，PAH患者可用于NO合成的精氨酸较少，继发于a）精氨酸酶活性增加，导致精氨酸向尿素和鸟氨酸的转化增加和2）精氨酸从头合成减少。 2.与健康对照组相比，PAH患者的NO合成较慢，因为不对称二甲基精氨酸（ADMA）抑制了一氧化氮合酶的活性。因此，肺动脉压和疾病严重程度将与NO合成速率呈负相关，与血浆ADMA浓度呈正相关。 二、具体目标 为了检验上述假设，将使用稳定同位素示踪技术和生化方法对3组受试者进行以下体内测量：特发性肺动脉高压患者、镰状细胞病和肺动脉高压患者以及对照组。 1.精氨酸通量（产生率或出现率）、其血浆浓度及其前体瓜氨酸从头合成精氨酸的速率。 2.尿素通量和鸟氨酸通量（精氨酸酶活性指标）、血浆鸟氨酸浓度以及精氨酸与鸟氨酸的血浆浓度比值 3.瓜氨酸通量、血浆浓度和精氨酸转化为瓜氨酸的速率（NO合成的指标） 4.不对称二甲基精氨酸的血浆浓度。 这些测量值将与血流动力学和临床参数相关，包括：肺动脉压、右心房压、超声心动图或右心导管检查的心脏指数和6分钟步行距离。

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. ABSTRACT Pulmonary arterial hypertension (PAH) is a disease of unknown etiology that is characterized by increased pulmonary artery pressure and vascular resistance. It can be idiopathic or secondary to other disease conditions, such as sickle cell disease. Decreased production of the vasodilatory molecule nitric oxide (NO) is believed to play an important role in the pathogenesis of PAH, but further in vivo studies of NO metabolism in patients with PAH are needed. We propose that there is a shortage of NO in PAH resulting from the combined effects of decreased availability of the NO precursor arginine and impaired activity of nitric oxide synthase, the enzyme that synthesizes NO. Specifically, we hypothesize that compared to healthy controls, patients with PAH will have less arginine available for NO synthesis secondary to increased activity of arginase, the enzyme that breaks down arginine to urea and ornithine, and decreased de novo arginine synthesis. In addition, they will have increased concentrations of asymmetric dimethylarginine, leading to inhibition of NO synthase activity. To test these hypotheses, stable isotope tracer techniques will be used to measure arginine and nitric oxide metabolism in healthy controls, patients with idiopathic PAH, and patients with PAH secondary to sickle cell disease. These measurements will be correlated with hemodynamic and clinical parameters. This project will increase our knowledge about the role of the arginine-NO pathway in the pathogenesis of PAH. In addition, it may identify future targets for therapeutic trials. I. HYPOTHESIS In this project, we propose to test the general hypothesis that decreased availability of the vasodilatory molecule nitric oxide (NO) underlies the pathogenesis of pulmonary arterial hypertension (PAH). This shortage of NO is a result of decreased synthesis due to the combined effects of the decreased availability of the NO precursor arginine and impaired nitric oxide synthase activity. Specific hypotheses to be tested are as follows: 1. Compared to healthy controls, patients with PAH have less arginine available for NO synthesis secondary to a) increased arginase activity leading to increased conversion of arginine to urea and ornithine and 2) decreased de novo arginine synthesis. 2. Compared to healthy controls, patients with PAH will have slower NO synthesis because of inhibition of nitric oxide synthase activity by asymmetric dimethylarginine (ADMA). As a consequence, pulmonary artery pressures and disease severity will be negatively correlated with the rate of NO synthesis and positively correlated with plasma ADMA concentration. II.SPECIFIC AIMS To test the above hypotheses, stable isotope tracer techniques and biochemical methods will be used to make the following measurements in vivo in 3 groups of subjects: patients with idiopathic pulmonary hypertension, patients with sickle cell disease and pulmonary hypertension, and controls. 1. Arginine flux (rate of production or rate of appearance), its plasma concentration, and the rate of de novo arginine synthesis from its precursor, citrulline. 2. Urea flux and ornithine flux (indices of arginase activity), plasma ornithine concentration, and the ratio of plasma concentration of arginine to ornithine 3. Citrulline flux, its plasma concentration, and the rate of conversion of arginine to citrulline (an index of NO synthesis) 4. Plasma concentration of asymmetric dimethylarginine. These measurements will be correlated with hemodynamic and clinical parameters, including: pulmonary artery pressures, right atrial pressure, and cardiac index by echocardiogram or right heart catheterization, and six minute walk distance.