REGULATING ION CHANNEL ACTIVITY IN CF AIRWAY EPITHELIA

调节 CF 气道上皮的离子通道活性

• 批准号: 3239434
• 负责人: Monroe JACKSON Stutts
• 金额: $ 8.5万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-03-01 至 1991-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3239434
• 关键词: autoradiography; autosomal recessive trait; cyclic AMP; cystic fibrosis; dogs; human tissue; immunochemistry; ion transport; phosphoproteins; phosphorylation; protein kinase; respiratory epithelium

In normal airway epithelial cells, chloride channels can be activated by agents which raise intracellular c-AMP. CF airway epithelial cells have been shown to contain chloride channels which cannot be activated by a normal increase in intracellular c-AMP. The defect in CF may involve some step in an activation sequence distal to the generation of c-AMP. Alternatively, CF cells could cause Cl-channel activity to be tonically inhibited. We propose to isolate and identify substances in the cytosol or associated with cell membranes which control the activity of single chloride channels in CF and non-CF airway epithelial cells. The specific aims of this proposal are: (1) Assay cytoplasm and membrane fractions derived from CF and non-CF cells for the ability to activate or deactivate chloride channels in the apical cell membrane of CF and non-CF cells and compare these effects to the effects of purified protein kinases or phosphatases. (2) Identify and isolate c-AMP dependent protein kinase activity, phosphatase activity, and phosphoproteins in cytoplasm and membrane fractions derived from CF and non-CF cells and assay for ability to alter ion channel function. The results from the proposed experiments should reveal if CF cells produce an inhibitor of ion channel activation, or if CF cells are missing a portion of the normal activation sequence of Cl channels found in normal cells.

在正常气道上皮细胞中，氯离子通道可以 由提高细胞内 c-AMP 的药物激活。 CF气道 上皮细胞已被证明含有氯离子通道 细胞内 c-AMP 的正常增加不能被激活。 CF 中的缺陷可能涉及激活序列中的某些步骤 远离 c-AMP 的产生。 或者，CF 细胞可以 导致 Cl 通道活性受到强直抑制。 我们建议 分离和鉴定细胞质或相关物质中的物质 具有控制单氯活性的细胞膜 CF 和非 CF 气道上皮细胞中的通道。 具体的 该提案的目的是： (1) 测定来自 CF 和的细胞质和膜组分 非 CF 细胞能够激活或停用氯离子 CF 和非 CF 细胞顶端细胞膜上的通道 将这些效果与纯化蛋白激酶的效果进行比较 或磷酸酶。 (2) 鉴定并分离 c-AMP 依赖性蛋白激酶活性， 磷酸酶活性以及细胞质和膜中的磷蛋白 来自 CF 和非 CF 细胞的级分和能力测定 改变离子通道功能。 拟议实验的结果应该揭示 CF 细胞是否 产生离子通道激活抑制剂，或者如果 CF 细胞 缺少 Cl 通道正常激活序列的一部分 存在于正常细胞中。