NMR STUDIES OF METAL IONS IN INTACT CELLS AND TISSUES

完整细胞和组织中金属离子的核磁共振研究

• 批准号: 3230494
• 负责人: Raj K Gupta
• 金额: $ 18.93万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1982
• 资助国家: 美国
• 起止时间: 1982-05-01 至 1989-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3230494
• 关键词: bleomycin; carbonate dehydratase; chemical structure function; conformation; cytochrome c; diphosphoglycerate; enzyme substrate complex; erythrocytes; heme; hemoglobin; hormone regulation /control mechanism; human tissue; hypertension; insulin; ion transport; lymphocyte; metalloproteins; nuclear magnetic resonance spectroscopy; oxyhemoglobin; phosphotransferases; sickle cell anemia

Intracellular metal ions modulate a variety of cellular and tissue functions. A long range goal of our research is to understand possible regulatory roles of these ions in cellular proliferation, differentiation, volume regulation, and hormonal control of various cellular processes. We seek also to understand how the concentrations of intracellular ions are managed by various membrane transport processes, and how such regulation goes astray in disorders such as cancer, hypertension, diabetes, and sickle cell disease. Our primary research tool is NMR spectroscopy. We have recently developed new NMR approaches for measuring free Mg++ (by analyzing 31P NMR of intracellular ATP) and Na+ ions (using an anionic paramagnetic reagent Dy(PPPi)2 introduced by us to distinguish between intra- and extracellular Na+ ions). The choice of cells and tissues for our research includes human normal and sickle erythrocytes, peripheral blood lymphocytes, isolated rat cardiac myocytes, amphibian eggs, and rat tail artery. Specific aims to be pursued are: (1) To determine if bound Na+ exists, and is functionally significant, by quantitating NMR-visible and total intracellular Na+ ion concentrations in various types of cells and physiological states; (2) If indeed a sizable part of intracellular Na+ in human erythrocytes is NMR-invisible as indicated by our preliminary experiments, to determine if it is sequestered in the cell membrane and cytoskeleton, and whether it can be released by intracellular Ca++ ions; (3) To study localization of Na+ ions in various intracellular compartments in amphibian oocytes; (5) To find out if partially relaxed 23Na FT NMR spectra can be used to detect the existence of compartments with differing relaxation behaviour and magnetic environments within an intact cell; (5) To follow changes in [Nai], pHi, [Mgi2+] during mitotic cell division cycles in synchronously dividing amphibian oocytes; (6) To ascertain if an abnormality in the regulation of intracellular Na+ ions is associated with the disordered mitotic rate and maturation sequence of leukemic lymphocytes; (7) To characterize the state of Na+ and K+ ions in normal and sickle red blood cells in oxygenated and deoxygenated states; (8) To investigate the effects of insulin upon membrane systems involved in the transport of Na+, K+ and H+ ions in amphibian oocytes and mammalian cardiac myocytes and to determine whether such effects are mediated by proteolytically generated peptides; and (9) To test the hypothesis that an increased NMR-visible intracellular Na+ is associated with essential hypertension.

细胞内金属离子调节多种细胞和组织 功能协调发展的 我们研究的一个长期目标是了解 这些离子在细胞增殖，分化， 体积调节和各种细胞过程的激素控制。 我们 也试图了解细胞内离子的浓度是如何 由各种膜运输过程管理，以及这种调节如何 在癌症、高血压、糖尿病和镰刀病等疾病中误入歧途 细胞疾病 我们的主要研究工具是NMR光谱。 我们有 最近开发了新的NMR方法用于测量游离Mg++（通过分析 细胞内ATP的31 P NMR）和Na+离子（使用阴离子顺磁共振（NMR））。 我们引进的Dy（PPPi）2试剂，用于区分细胞内 细胞外Na+离子）。 我们研究的细胞和组织的选择 包括人正常和镰状红细胞、外周血 淋巴细胞、离体大鼠心肌细胞、两栖类卵和大鼠尾 动脉 具体目标是：（1）确定结合Na+ 存在，并具有功能意义，通过定量NMR可见， 各种类型细胞中的总细胞内Na+离子浓度， 生理状态;（2）如果细胞内Na+确实有相当大的一部分， 我们的初步研究表明，人红细胞是NMR不可见的。 实验，以确定它是否被隔离在细胞膜中， 细胞骨架，以及它是否能被细胞内Ca++离子释放; (3)研究Na+离子在细胞内不同区室中的定位 （5）部分弛豫的~（23）Na FT NMR 光谱可用于检测具有不同的 完整细胞内的弛豫行为和磁环境;（5） 观察细胞分裂过程中[Nai]、pHi、[Mgi 2 +]的变化 两栖类卵母细胞同步分裂的周期;（6）为了确定 细胞内Na+离子调节的异常与 白血病细胞有丝分裂率和成熟顺序紊乱 （7）测定正常人和正常人外周血淋巴细胞中Na ~+和K ~+离子的状态， 镰状红细胞处于氧合和脱氧状态;（8） 研究胰岛素对膜系统的影响， Na ~+、K ~+和H ~+离子在两栖类卵母细胞和哺乳动物心脏中的转运 并确定这种作用是否由 蛋白水解产生的肽;和（9）为了测试假设， 增加的NMR可见细胞内Na+与必需的 高血压