MODEL FOR GLUCOSE INDUCED BASEMENT MEMBRANE THICKENING

葡萄糖引起的基底膜增厚模型

• 批准号: 2444097
• 负责人: MICHAEL Peter SARRAS
• 金额: $ 18.76万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-07-01 至 1999-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2444097
• 关键词: Cnidaria; SDS polyacrylamide gel electrophoresis; alternatives to animals in research; basement membrane; cell cell interaction; cellular pathology; collagen; enzyme activity; epithelium; extracellular matrix; fibronectins; genetic library; genetic translation; immunocytochemistry; laminin; medical complication; membrane biogenesis; membrane structure; messenger RNA; metalloendopeptidases; model; molecular pathology; monoclonal antibody; nucleic acid probes; polymerization; western blottings

A thickening in basement membranes (or epithelial-associated extracellular matrix (ECM)) is a common feature of a number of disease states. For example, ECM thickening has been observed 1) associated with the renal tubules and glomeruli (GBM) of patients with polycystic kidney disease (PKD), 2) associated with the seminiferous tubules in male patients with certain forms of impaired fertility, 3) associated with the GBM of patients with Alport syndrome, and 4) in patients with diabetes mellitus. Progressive secondary complications of these diseases have been tied to abnormalities associated with this ECM thickening. Although an explanation as to the mechanisms by which abnormal ECM thickening occurs is not currently available, it is evident that the normal biosynthesis and maintenance of ECM is an important aspect of healthy differentiated tissues. As indicated, the cellular and molecular basis for alterations in ECM formation is not understood, but may be explained in terms of problems in the balance of ECM component synthesis, polymerization, and turnover. These processes all involve cell/ECM interactions to some degree and imply that ECM formation is normally tightly controlled by both cellular and extracellular processes. Experimental approaches to evaluate the mechanisms which govern normal ECM formation and the mechanisms by which abnormally thickened ECM occurs have been hampered by a lack of in vitro and in vivo cellular models. In order to approach these problems we have developed an in vivo cellular model in which epithelial-associated ECM formation can be experimentally induced in a short time frame for subsequent analysis of the cellular mechanisms involved in the process. In addition, methods have been developed to trigger the formation of abnormally thickened ECM so that both the normal and abnormal process can be directly compared and evaluated. The model we have developed is simply comprised of an epithelial bilayer with an intervening ECM. In addition, we have determined that this in vivo model responds to hyperglycemic conditions by thickening its ECM as observed in various pathological conditions. As opposed to vertebrate animal models currently available however, the cell system we utilize develops an ECM within 24-96 hr and doubles the thickness of this ECM within this same time frame when exposed to elevated levels of glucose. This model was developed using the Cnidarian, Hydra vulgaris. The proposed project will utilize this in vivo model to analyze normal ECM formation and determine if abnormal thickening of ECM results from 1) cellular abnormalities in the synthesis and accumulation of ECM components, 2) abnormalities in the extracellular assembly of ECM components., and/or 3) abnormalities in the degradation of ECM components. A combination of morphological, biochemical, and molecular approaches will be utilized to test these hypotheses. This project will provide basic information on the cellular mechanisms of ECM formation under normal conditions and under conditions in which ECM thickening occurs.

基底膜(或上皮相关的细胞外)增厚 矩阵(ECM))是许多疾病状态的共同特征。为 例如，已观察到1)与肾脏相关的ECM增厚 多囊肾病患者肾小管和肾小球的变化 (PKD)，2)与男性患者的生精小管相关 某些形式的生育能力受损，3)与 Alport综合征患者，以及4)糖尿病患者。 这些疾病的进行性继发性并发症与 与此ECM增厚相关的异常。尽管有一个 细胞外基质异常增厚发生机制的解释 目前还不可用，很明显，正常的生物合成和 细胞外基质的维持是健康分化的重要方面 纸巾。正如所指出的，改变的细胞和分子基础 在ECM的形成是不了解的，但可以解释为 ECM组分合成、聚合和平衡中存在的问题 营业额。这些过程都涉及到细胞/细胞外基质的相互作用 程度并暗示ECM的形成通常由两者严格控制 细胞和细胞外过程。评估的实验方法 控制正常细胞外基质形成的机制和通过 发生的异常增厚的ECM由于缺乏 体外和体内细胞模型。为了解决这些问题，我们 已经开发出一种体内细胞模型，在这种模型中，上皮细胞与 ECM的形成可以在很短的时间内被实验诱导 随后对该过程中涉及的细胞机制进行分析。 此外，已经开发了一些方法来触发 异常增厚的ECM，因此正常和异常过程都可以 被直接比较和评价。我们开发的模型很简单 由具有介入性ECM的上皮性双层组成。此外, 我们已经确定这个体内模型对高血糖有反应 条件通过增厚其ECM在不同的病理观察 条件。与目前可用的脊椎动物模型相反 然而，我们使用的细胞系统在24-96小时内产生ECM，并且 在相同的时间范围内将此ECM的厚度增加一倍 导致血糖水平升高。此模型是使用 刺蛇，九头蛇。拟议的项目将在体内利用这一点。 分析正常ECM形成并确定是否异常增厚的模型 ECM的结果来自1)细胞合成和 细胞外基质成分的积累，2)细胞外的异常 ECM组件的组装，和/或3)降解异常 ECM组件。形态、生化和 分子方法将被用来检验这些假说。这 该项目将提供有关ECM细胞机制的基本信息 在正常条件下和在ECM条件下形成 就会发生增厚。