OXIDATION AND DEFORMATION IN SICKLE RBC DEHYDRATION

描述：此修订后的续期申请的目的是 提高我们对分子机制的理解， 镰状红细胞异常脱水，可以说是最 这些细胞的重要细胞缺陷，并进一步阐明 铁和氧化在镰状细胞病发病中的作用 疾病该提案包括两个分项目， 因为它们都与阳离子的问题有关， 体内平衡和氧化病理学。 的作用研究 氧化在镰状病病理生理学中具有三个特异性 目的：1）确定增加“自动氧化”的原因， 镰状血红蛋白，考虑到它不是来自固有的 自氧化不稳定性，而不是从 镰刀型相互作用 血红蛋白与过渡金属，病理生理氧化剂， 膜磷脂; 2）定义铁的性质 通过评估镰状红细胞中的去部分化：细胞质游离 过渡金属含量及其与无膜铁的关系， 以及铁的平衡结合， 细胞溶质生理螯合剂;金属/血红蛋白/膜的作用 膜铁沉积中的相互作用;和铁隔室， 基因型的函数; 3）测试关于生物化学的预测 这种氧化过程的后果，包括验证 假设： 不稳定的血红蛋白病膜是镰状的 以及使用铁螯合剂去除红细胞膜铁 应该会带来可预测的改善，这种方法将测试 一种潜在的治疗选择 膜作用的研究 变形诱导的红细胞阳离子泄漏具有三种特异性 （1）界定解释《公约》独特特征的机制， 低渗性渗漏 变形 领导 假设 是 测试的是，大分子拥挤发挥了主要作用， 变形对膜响应的影响; 2)建立 这个模型的关系，以真实的镰刀通过测试 预测镰状细胞的质量特征 泄漏将取决于变形的程度;详细介绍其比较 现象学;并确定其与膜脂质的关系 氢过氧化物; （3）关于红色是否 细胞机械敏感性发生的设计或意外，通过检查 已知泄漏途径的选定红细胞的变形反应 缺乏;并通过使用锌处理的红细胞来检查 假设允许机械敏感泄漏结构 由带3蛋白聚集而成。 预计各国 成功实现这些目标将有助于 在我们的分子基础的理解显着改善， 镰状红细胞膜病理学和细胞脱水， 也为镰状细胞病带来了新的治疗方法。

DESCRIPTION: The objective of this revised renewal application is to improve our understanding of the molecular mechanisms responsible for abnormal dehydration of sickle red cells, arguably the most important cellular defect of these cells and further clarifying the role of iron and oxidation as pathogenetic influences in sickle cell disease. The proposal comprises of two subprojects that are interrelated in that they both are relevant to problems of cation homeostasis and of oxidative pathobiology. The study on the role of oxidation in sickle disease pathophysiology has three specific aims: 1) Identify the reason for increased "autoxidation" of sickle hemoglobin, considering that it derives not from an inherent autoxidative instability but rather from interaction of sickle hemoglobin with transition metals, pathophysiologic oxidants, and membrane phospholipid; 2) Define the nature of iron decompartmentalization in sickle red cells by assessing: cytoplasmic free transition metal content and its relationship to membrane-free iron, as well as equilibrium binding of iron as influenced by cytosolic physiologic chelators; the role of metal/hemoglobin/membrane interaction in membrane iron deposition; and iron compartments as a function of genotype; 3) Test predictions about the biochemical consequences of such oxidative processes including the validating of hypotheses: that unstable hemoglobinopathy membranes are sickle- like; and that removal of red cell membrane iron using an iron chelator should effect predictable improvements, an approach that will test a potential therapeutic option. The study on the role of membrane deformation-induced cation leak from red cells has three specific aims: 1) Define mechanisms explaining the unique features of the leak resulting from hypotonic deformation. The leading hypothesis to be tested is that macromolecular crowding exerts a major influence on membrane's response to deformation; 2) Establish the relationship of this model to authentic sickling by testing the prediction that the qualitative character of the sickling-induced leak will depend on the degree of deformation; detailing its comparative phenomenology; and determining its relationship to membrane lipid hydroperoxides; and 3) Approach the question as to whether red cell mechanosensitivity occurs by design or accident by examining the deformation response of selected red cells with known leak pathway deficiencies; and by using zinc treated red cells to examine the hypothesis that a permissive mechanosensitive leak structure is formed by clustering of Band 3 protein. It is anticipated that successful accomplishment of these objectives will contribute to a significant improvement in our understanding of the molecular basis for sickle red cell membrane pathobiology and cell dehydration and could also lead to novel therapeutic approaches to sickle cell disease.