DEVELOPMENT AND MAINTENANCE OF EPITHELIAL CELL POLARITY

上皮细胞极性的发育和维持

基本信息

批准号：
3086915
负责人：
PHILIP P BREITFELD
金额：
$ 7.65万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
1986
资助国家：
美国
起止时间：
1986-07-01 至 1991-06-30
项目状态：
已结题

项目摘要

The mechanisms by which organs and their component cells acquire differentiated functions and morphology are critical to normal human development. This functional and morphologic regionality of cells is referred to as polarity. Epithelial cells, and in particular hepatocytes, serve as a useful model to study the development and maintenance of cell polarity since hepatocytes display morphologic polarity in monolayer culture. In addition, they are developmently regulated to perform many metabolic functions and display biochemical markers for human development (e.g. alpha fetoprotein). To examine the mechanisms whereby epithelial cells develop and maintain polarity, human hepatoma-derived cell lines will be established in monolayer culture on suspended Millipore filters. The formation of an intact, polarized monolayer will be assessed by electron microscopy and determination of electrical resistance across the monolayer. Using this culture system, the organization of intracellular organelles and cell-cell junctions will be assessed by electron microscopy. The localization of cell nutrient and growth factor entry will be determined by radio-ligand binding assay and by localization of the growth factor receptors using immunofluorescence. The intracellular traffic of nutrients and growth factors as well as their receptors will be examined using the asialoglycoprotein (ASGP) receptor system as a model. The fate of both the ligand (ASGP) and the receptor will be determined biochemically. The development of cell-cell communication and cell adhesion will be addressed by determining the localization of cell adhesion molecules in human hepatoma-derived cell lines. In addition, the role of tight junctions in maintenance of surface membrane polarity will be assessed by determining the fate of polarized integral membrane proteins after disruption of tight junctions. The development of specialized membrane domains will be examined in this system by determining the steady state localization of individual integral membrane proteins as well as their time of appearance at the cell surface following biosynthesis. Finally, the rates and sites of secretion for a host of secretory proteins will be determined. Thus, this system will allow for the direct examination of several aspects of cell polarity which will aid in understanding normal morphogenesis, embryogenesis, and fetal development.

器官及其组成细胞获得的机制分化的功能和形态对正常人至关重要发展。细胞的这种功能和形态区域性是称为极性。上皮细胞，特别是肝细胞，作为研究细胞发育和维持的有用模型极性，因为肝细胞在单层中表现出形态极性文化。此外，它们受到发展监管以执行许多代谢功能和显示人类发育的生化标记（例如甲胎蛋白）。研究上皮细胞发育和维持的机制极性，人肝癌细胞系将在在悬浮微孔过滤器上进行单层培养。的形成完整的偏振单层将通过电子显微镜进行评估测定跨单层的电阻。使用这种培养系统，细胞内细胞器的组织和细胞与细胞的连接将通过电子显微镜进行评估。这细胞营养和生长因子进入的定位将取决于放射性配体结合测定和生长因子的定位使用免疫荧光的受体。细胞内营养物质的运输生长因子及其受体将使用以下方法进行检查脱唾液酸糖蛋白（ASGP）受体系统作为模型。两人的命运配体（ASGP）和受体将通过生化方法测定。细胞间通讯和细胞粘附的发展将通过确定细胞粘附分子的定位来解决人肝癌来源的细胞系。另外，还有紧缩的作用维持表面膜极性的连接将通过以下方式评估确定极化后整合膜蛋白的命运紧密连接的破坏。本研究将研究专门膜域的发展通过确定个体积分的稳态定位来系统膜蛋白及其在细胞表面出现的时间生物合成后。最后，分泌速率和分泌位点分泌蛋白的宿主将被确定。因此，该系统将允许直接检查几个方面细胞极性这将有助于理解正常的形态发生，胚胎发生和胎儿发育。