Role of hemoglobin b93cys residue in nitric oxide bioactivity

描述（由申请人提供）：血红蛋白293 cys残基是保守的，最近作为红细胞如何影响血管一氧化氮（NO）代谢和功能的潜在调节剂受到了广泛关注。然而，确切的功能和机制仍不清楚。了解这种残留物如何控制血管一氧化氮功能是至关重要的，因为这些机制的功能障碍可能导致许多血管病理状态。在这个建议中，我们建立在从新的小鼠模型产生的初步数据上，这些小鼠模型专门表达野生型人血红蛋白或人血红蛋白，其中293 cys残基在其红细胞中被Ala取代。具体而言，我们目前的数据表明，在生理条件下，脱氧的红细胞和血红蛋白激活亚硝酸还原酶的活性，导致在一个电子还原的阴离子亚硝酸盐NO。这个过程是通过控制血红蛋白的氧亲和力和有趣的调制293 cys残基变构调节。在这种情况下，我们提出，293半胱氨酸残基是至关重要的耦合血红蛋白氧传感与亚硝酸盐衍生NO-生物活性。相反，在急性炎症性疾病脓毒症期间，我们提出293 cys是亚硝化应激形成S-亚硝基血红蛋白（SNOHb）的靶点，这反过来又导致与这种疾病相关的血管和肺功能障碍。后者由显示含SNO的红细胞可以刺激中性粒细胞粘附到肺内皮细胞并以不依赖于变构调节的方式引起血管舒张的数据指示。肺部炎症和肺损伤都是脓毒症的特征。这些新概念将在本提案中通过追求以下假设来研究：在急性炎症期间，293 cys残基作为血管NO信号传导的调节剂的作用从亚硝酸盐还原酶依赖性机制改变为SNOHb依赖性机制，这将通过以下具体目的进行测试：1）确定293 cys调节RBC依赖性NO血管细胞信号传导的机制。2)确定293 cys残基在体内控制亚硝酸盐还原和NO依赖性血管细胞信号传导中的作用。3）确定293 cys残基在脓毒症诱导的低血压和肺部炎症期间影响RBC效应中的作用。这些目标的实现将有助于深入了解红细胞如何调节NO代谢的新的治疗靶点的机制。

DESCRIPTION (provided by applicant): The hemoglobin 293cys residue is conserved and received much attention recently as a potential modulator of how red cells affect vascular nitric oxide (NO) metabolism and function. However, the precise function and mechanisms involved remain unclear. Understanding how this residue controls vascular nitric oxide function is critical since dysfunction in these mechanisms may contribute to a number of vascular pathological states. In this proposal we build upon preliminary data generated from novel mouse models that express exclusively either wild-type human hemoglobin or human hemoglobin in which the 293cys residue has been replaced with an Ala in their red cells. Specifically, we present data indicating that under physiological conditions, deoxygenation of red cells and hemoglobin activates a nitrite reductase activity that results in the one-electron reduction of the anion nitrite to NO. This process is regulated allosterically by controlling hemoglobin oxygen affinity and interestingly modulated by the 293cys residue. In this context we propose that the 293cys residue is critical in coupling hemoglobin oxygen sensing with nitrite derived NO-bioactivity. In contrast, during the acute inflammatory disease Sepsis, we propose that the 293cys is a target for nitrosative stress forming S-nitrosohemoglobin (SNOHb), which in turn contributes to the vascular and pulmonary dysfunction associated with this disease. The latter is indicated by data showing SNO-containing red cells can stimulate neutrophil adhesion to pulmonary endothelial cells and elicit vasodilation in a manner that is independent on allosteric regulation. Both pulmonary inflammation and lung injury are features of sepsis. These novel concepts will be investigated in this proposal by pursuit of the hypothesis that during acute inflammation, the role of the 293cys residue as a modulator of vascular NO-signaling changes from a nitrite-reductase dependent to SNOHb dependent mechanism which will be tested via the following specific aims 1) Determine the mechanism by which 293cys regulates RBC dependent NO vascular cell signaling., 2) Determine the role of 293cys residue in controlling nitrite reduction and NO- dependent vascular cell signaling in vivo, 3) Determine the role of the 293cys residue in affecting RBC effects during Sepsis induced hypotension and pulmonary inflammation. Accomplishment of these aims will yield insights into the mechanisms novel therapeutic targets focusing on how RBCs modulate NO-metabolism.