PEROXYNITRITE AND NITROTYROSINE IN HEART DISEASE

描述：扩散限制反应生成的过氧亚硝酸根(ONOO-) 超氧化物歧化与一氧化氮结合是脑组织氧化损伤的主要机制 心脏。暴露于微摩尔浓度的离体心肌 过氧亚硝酸盐仅在几分钟内就表现出许多病理性 临床心脏病的特征变化，包括损伤到 线粒体、收缩功能丧失和不可逆转的增加 舒张期僵硬。过氧亚硝酸盐攻击的主要后果之一 是特定蛋白质中酪氨酸残基的选择性硝化。 大鼠心肌细胞在缺血/再灌流过程中迅速硝化 抑制一氧化氮的产生既能保护心肌功能 并减少酪氨酸硝化。在人类的心脏中，类似的模式 心肌细胞硝化发生在缺血、脓毒症和心肌炎之后。 定量高效液相分析显示，1-7%的蛋白质结合酪氨酸是 败血症患者的硝化作用，表明硝化作用是一种重要的病理变化 在人类的过程中。西方印迹显示令人惊讶的是有限的 一些蛋白质容易受到过氧亚硝酸盐对酪氨酸硝化的影响。 首席研究人员表明，锰超氧化物歧化酶， 肌动蛋白和中间丝在人体尸检材料中硝化 这些蛋白质的功能被过氧亚硝酸根显著削弱 在试管中。电喷雾质谱仪显示，过氧亚硝酸盐 优先硝化亲水性区域中的某些酪氨酸基序 为许多结构蛋白所共有的。首席调查者提出假设 过亚硝酸根对结构蛋白的酪氨酸硝化作用 到心肌损伤。他的第一个目标是定量评估 大鼠心肌缺血后酪氨酸亚硝化量的变化 使用他们开发的一种新型探头来测量过亚硝酸根。第二 目的是确定在大鼠和人类中哪些特定的蛋白质是硝化的 心脏，并用串联质谱仪测定这些物质中的哪些酪氨酸 蛋白质被硝化。第三个目标是描述硝化作用如何 然后对影响肌动蛋白功能的其他结构蛋白进行评估 硝化作用如何影响肌肉收缩蛋白。这些研究是一项新的 和新的方法来理解具体的机制 生物产生的氧化剂会损害心脏，并使我们能够识别 体内过氧亚硝酸盐攻击的主要目标。

DESCRIPTION: Peroxynitrite (ONOO-) formed by the diffusion-limited reaction of superoxide with nitric oxide is a major mechanism of oxidative damage in the heart. Isolated heart muscle exposed to micromolar concentrations of peroxynitrite for only a few minutes exhibit many of the pathological changes characteristic of clinical heart disease, including injury to mitochondria, loss of contractile function and irreversible increases in diastolic stiffness. One of the major consequences of peroxynitrite attack is the selective nitration of tyrosine residues in specific proteins. Myocytes in rat heart become nitrated quickly during ischemia/reperfusion and inhibition of nitric oxide production both preserves myocardial function and reduces tyrosine nitration. In the human heart, similar patterns of myocyte nitration occur after ischemia, sepsis and myocarditis. Quantitative HPLC analysis revealed 1-7% of protein-bound tyrosines are nitrated in septic patients, showing nitration is a significant pathological process in humans. Western blots demonstrate that a surprisingly limited number of proteins are vulnerable to tyrosine nitration by peroxynitrite. The principal investigator has shown that manganese superoxide dismutase, actin and intermediate filaments are nitrated in human autopsy material and that the function of these proteins is markedly diminished by peroxynitrite in vitro. Electrospray mass spectrometry has revealed that peroxynitrite preferentially nitrates certain tyrosine motifs in hydrophilic regions common to many structural proteins. The principal investigator hypothesizes that tyrosine nitration of structural proteins by peroxynitrite contributes to myocardial injury. His first aim is to quantitatively assess the magnitude of tyrosine nitrosation in ischemic rat heart and to directly measure peroxynitrite using a novel probe they have developed. The second aim is to identify which specific proteins are nitrated in rat and human heart and determine by tandem mass spectrometry which tyrosines in these proteins are nitrated. The third aim is to characterize how nitration affects the function of actin and then other structural proteins to assess how nitration can affect myocontractile proteins. These studies are a new and novel approach to understanding the specific mechanisms by which biologically generated oxidants injure the heart and allow us to identify major targets for peroxynitrite attack in vivo.