MEMBRANE PERTURBATION DURING INFLAMMATION

这项建议是对最近的小额赠款计划的回应 训练有素、经验较少的基础科学家。具体目标和 长期目标是开发牛角膜杯作为一种治疗 研究炎症过程中膜的变化。这种方法 将允许剥离炎症细胞--角膜内皮细胞 从形态和功能的观点来看，相互作用。许多 眼睛内炎症过程的分支最终可能是 通过使用特定的细胞类型、受刺激的细胞(即，激活的 中性粒细胞、淋巴细胞、巨噬细胞)、特异性体液或化学物质 炎症、病毒或细菌疾病状态的介质，以及阻断 凝集素等受体。 在这项初步研究中将解决几个具体问题：1) 角膜内皮细胞表面形态发生了什么变化？ 炎症细胞黏附过程中的糖类结构 轮回？；2)轮回的形态或功能有变化吗？ 内皮细胞间连接？以及3)这些变化是否依赖于 一种特殊的炎性细胞类型？ 为了开始回答这些问题，牛角膜杯模型 将通过将几种技术应用于该模型来建立。 将分离特定的外周血白细胞。中的更改 内皮细胞表面形态将通过使用 扫描电子显微镜和冷冻断裂技术 膜内颗粒测定积累的定量数据 大小和密度分布。细胞表面碳水化合物的变化 结构将通过一组凝集素与之复合而确定 然后可以使用扫描电子来定量血蓝蛋白 显微镜。细胞间连接的变化将通过以下方式确定 对交叉口大小和分布的定量评估 冷冻断裂技术。 角膜杯模型的建立将允许更可控和 详细检查炎症细胞与内皮细胞的相互作用。

This proposal is in response to the small grants program for recently trained, less experienced basic scientists. The specific aims and long-term objectives are to develop the bovine corneal cup as a method for studying changes in membrane during the inflammatory process. This method will allow dissection of the inflammatory cell-corneal endothelial cell interaction from a morphologic and functional viewpoint. Many ramifications of the inflammatory process within the eye can eventually be explored by use of specific cell types, stimulated cells (i.e., activated neutrophils, lymphocytes, macrophages), specific humoral or chemical mediators of inflammation, viral or bacterial disease states, and blocking receptors such as lectins. Several specific questions will be addressed in this initial study: 1) What are the changes in corneal endothelial cell surface morphology and carbohydrate structure during inflammatory cell adhesion and transmigration?; 2) Are there changes in the morphology or function of endothelial intercellular junctions?; and 3) Are these changes dependent on a specific inflammatory cell type? In order to begin answering these questions the bovine corneal cup model will be established with several techniques being applied to the model. Specific peripheral blood leukocytes will be isolated. Changes in endothelial cell surface morphology will be determined by the use of scanning electron microscopy and the freeze-fracture technique with quantitative data accumulated by determination of intramembranous particle size and density distribution. Changes in cell surface carbohydrate structure will be determined with a battery of lectins complexed with hemocyanin which then can be quantitated by use of scanning electron microscopy. Changes in intercellular junctions will be identified by quantitative assessment of junction size and distribution by use of the freeze-fracture technique. Establishment of the corneal cup model will allow a more controlled and detailed examination of inflammatory cell-endothelial cell interaction.