HETEROGENEITY CELL CALCIUM DISTRIBUTION RENAL HYPOXIA

异质性细胞钙分布肾缺氧

• 批准号: 3236753
• 负责人: EDORIS ANN LEFURGEY
• 金额: $ 14.45万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-04-15 至 1991-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3236753
• 关键词: acidity /alkalinity; adenine nucleotides; calcium metabolism; cell death; homeostasis; hypoxia; kidney circulation; laboratory rabbit; mannitol; mitochondria; organelles; renal ischemia /hypoxia; renal tubular transport

Our long range goal is to define the role which calcium plays in early events leading to renal ischemic cell injury. To do so, it is vital to identify the sites of release, uptake and binding of calcium within the cells. Electron probe x-ray microanalysis (EPXMA) offers and unexcelled means of measuring elemental contents within individual cells and subcellular organelles, so that changes in the amounts of localized calcium can be quantitated at chosen intervals during the time course leading to irreversible cell injury. Preliminary experiments using EPXMA in proximal tubules have demonstrated a large degree of heterogeneity in the cellular and intracellular distribution of calcium. Such heterogeneity brings into question numerous previous studies which have assumed that there is a more homogeneous distribution. The proximal tubule suspension facilitates experimental maneuvers which permit examination of the earliest events leading to irreversible cell damage and eliminates secondary effects that, in an intact animal model, arise from compromising the function of the renovascular system in toto. We shall perform, in parallel, experiments which will measure biochemical, physiological, ultrastructural and microchemical variables on the same suspension. Experimental conditions will also be examined that preserve renal function during anoxia, such as the presence of adenine nucleotides, mannitol, and low pH in the bathing medium. Experiments will be performed to measure cytosolic free calcium using methyl ester dyes (fura-2) under identical conditions. EPXMA will be used to validate these measurement by determining the compartmentation of the dyes. This powerful technique (EPXMA) judiciously applied to a well-characterized renal preparation, in combination with the established array of physiological, biochemical, and ultrastructural measurements, is expected to provide unique direct information on the role of calcium in hypoxic cell injury in the kidney.

我们的长期目标是确定钙在 导致肾缺血性细胞损伤的早期事件。 为此， 关键是确定释放，吸收和结合的网站， 细胞内的钙。 电子探针x射线显微分析 （EPXMA）提供了测量元素的无与伦比的手段 在单个细胞和亚细胞器内的内容物，因此， 局部钙的量的变化可以定量， 在导致不可逆电池的时间过程中选择的间隔 损伤 在近端小管中使用EPXMA的初步实验 已经证明了在细胞中的很大程度的异质性， 和钙的细胞内分布。 这种异质性 这对之前的许多研究提出了质疑， 假设有一个更均匀的分布。 近端小管悬浮液有助于实验 允许检查最早事件的演习 导致不可逆的细胞损伤， 在一个完整的动物模型中， 肾血管系统的功能。 我们将 并行地进行实验，这些实验将测量生化， 生理、超微结构和微化学变量 同样的暂停。 还将检查实验条件 在缺氧期间保护肾功能，例如 腺嘌呤核苷酸，甘露醇，和低pH值的洗澡 介质 将进行实验以测量细胞溶质 游离钙，使用甲酯染料（Fura-2），在相同条件下 条件 EPXMA将用于验证这些测量结果 通过测定染料的区室化。 这个强大 技术（EPXMA）明智地适用于一个良好的特点， 肾脏准备，结合已建立的 生理、生化和超微结构测量， 预计将提供关于以下方面作用的独特直接信息： 肾脏缺氧细胞损伤中的钙。