CELL BIOLOGICAL STUDIES ON DIABETES AND METABOLISM

糖尿病和代谢的细胞生物学研究

• 批准号: 2138093
• 负责人: Leonard Jarett
• 金额: $ 28.18万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-01-01 至 1996-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2138093
• 关键词: cell cycle; cell growth regulation; cell nucleus; chemical aggregate; diabetes mellitus; electron microscopy; endocytosis; gene expression; genetic transcription; gold; hormone regulation /control mechanism; immunoelectron microscopy; in situ hybridization; insulin; insulin receptor; insulin sensitivity /resistance; insulinlike growth factor; intracellular transport; laboratory mouse; laboratory rat; liver; macromolecule; messenger RNA; monensin; muscle; mutant; northern blottings; peptide hormone metabolism; perfusion; receptor binding; thymidine; tissue /cell culture; tritium

The ultrastructural and biochemical studies in this project will increase our knowledge of the mechanisms that regulate the aggregation, internalization and intracellular processing of the insulin-receptor complex and will determine whether those processes, especially nuclear translocation of insulin and insulin's effects on macromolecular uptake into and efflux from the nucleus, are involved in insulin's regulation of cellular metabolism and growth. The specific aims for the next five years are: 1) To use immunoelectron microscopic techniques to document the organization, distribution, lateral mobility, internalization routes and intracellular processing of insulin receptors on liver and skeletal muscle. These tissues, in addition to the adipocyte that has already been studied, are the primary insulin-sensitive tissues. The three tissues differ in their responses to insulin. Tissue-specific differences in insulin-receptor complex processing, which may be observed, should be valuable in understanding the relationship between hormone-receptor complex processing and insulin action. 2) To use high resolution ultrastructural techniques and cells expressing either normal or mutated human insulin receptors to determine which domains of the insulin receptor are responsible for the normal organization, distribution, lateral mobility, internalization and intracellular processing of the insulin-receptor complex. These studies will provide ultrastructural analysis to complement biochemical data concerning the relationships between the molecular structure of the insulin receptor and the aggregation, internalization and appropriate intracellular targeting of the insulin-receptor complex leading to normal insulin action. Similar studies will be performed using cells expressing normal or mutated human IGF I receptors. 3) To use ultrastructural and biochemical techniques to characterize the intracellular route responsible for the translocation and nuclear accumulation of insulin. By determining this intracellular pathway, we should learn more about the mechanism by which insulin regulates gene transcription and cell growth. In some of these studies agents that affect the nuclear accumulation of insulin will be used to accentuate key components of the intracellular translocation itinerary. 4) To determine whether or not insulin's effects on cell proliferation or gene expression correlate with the nuclear accumulation of insulin and/or insulin's effects on macromolecular uptake into and efflux from the nucleus. Our hypothesis, that the nuclear accumulation of insulin and its effects on macromolecular translocation are physiologically significant and possibly related to cell growth and/or gene expression, will be tested by evaluating these processes in cell types, including those expressing mutated human insulin receptors, with different growth-related responses to insulin. Ultrastructural, cellular and molecular techniques, including in situ electron microscopic hybridization, Northern blot analysis, etc., will be used for these studies. We will determine whether insulin accumulation and insulin-stimulated macromolecular nuclear uptake is cell-cycle dependent. The information gained from studies in this project will provide insights into normal mechanisms of insulin action and potential causes of insulin-resistance and diabetes mellitus. These insights may then provide possible alternatives for intervention or therapy.

该项目的超微结构和生化研究将 提高我们对调节聚合的机制的了解， 胰岛素受体的内在化和细胞内处理 复杂并将确定这些过程是否，尤其是核 胰岛素和胰岛素对大分子摄取的影响的易位 核的进入和外排参与胰岛素调节 细胞代谢和生长。 接下来五个的具体目标 年是：1）使用免疫电子微观技术记录 组织，分销，横向移动性，内部化路线 以及肝脏和骨骼上胰岛素受体的细胞内加工 肌肉。 这些组织，除了已经具有的脂肪细胞 被研究为主要的胰岛素敏感组织。 三个 组织对胰岛素的反应不同。 组织特异性 胰岛素受体复合物处理的差异，这可能是 观察到的，对于理解之间的关系应该很有价值 激素受体复合物加工和胰岛素作用。 2）使用高 分辨率超微结构技术和表达正常的细胞 或突变的人类胰岛素受体，以确定哪些域 胰岛素受体负责正常组织， 分布，横向迁移率，内在化和细胞内 胰岛素受体复合物的处理。 这些研究将提供 超微结构分析以补充有关生化数据 胰岛素受体的分子结构之间的关系 以及聚集，内在化和适当的细胞内 靶向胰岛素受体复合物导致正常胰岛素 行动。 使用表达正常的细胞进行类似的研究 或突变的人IGF I受体。 3）使用超微结构和 生化技术以表征细胞内途径 负责胰岛素的易位和核积累。 通过确定这种细胞内途径，我们应该了解更多有关 胰岛素调节基因转录和细胞的机制 生长。 在某些影响核的研究代理中 胰岛素的积累将用于强调关键组成部分 细胞内易位行程。 4）确定是否 胰岛素对细胞增殖或基因表达相关的影响 随着胰岛素和/或胰岛素对核积累的影响 大分子从细胞核中摄取和排出。 我们的假设， 胰岛素的核积累及其对 大分子易位具有生理意义，并且 可能与细胞生长和/或基因表达有关，将测试 通过在细胞类型中评估这些过程，包括表达的过程 突变的人类胰岛素受体，具有不同的生长相关反应 胰岛素。 超微结构，细胞和分子技术， 包括原位电子微观杂交，北印迹 分析等将用于这些研究。 我们将确定 胰岛素积累和胰岛素刺激的大分子是否存在 核摄取是细胞周期依赖性的。 从中获得的信息 该项目的研究将提供有关正常机制的见解 胰岛素耐药和糖尿病的胰岛素作用和潜在原因 Mellitus。 这些见解可能会为 干预或治疗。