MECHANISM OF AIR POLLUTANT INDUCED AIRWAY PERMEABILITY

拟议的研究将调查 a) 控制气道的机制 渗透性，b) 吸入后早期损伤的细胞内部位 大鼠暴露于 03 和 N02 以及 c) 阻止或逆转污染物的方法 导致渗透性增加。 我们实验室的试点研究表明，气管和 吸入后支气管肺泡对大分子的通透性 将大鼠暴露于 03。似乎导致渗透性增加，在 至少部分是由于紧密连接的扰动。 基于我们的 肌动蛋白和肌球蛋白在体内分布的初步观察 顶端细胞质和纤毛紧密连接附近 上皮细胞，也基于行为的相似性 污染物气体和细胞骨架破坏药物，我们建议 细胞骨架可能是污染物的细胞内靶位点 它在紧密连接渗透性的调节中起着关键作用。 我们还预计细胞骨架在囊泡运输中的作用 与其他系统一样的气道。 拟议的研究考虑到 细胞骨架活性药物与污染物气体的广泛比较 药物可能逆转污染物的影响。 各种 将分析指标以实现预计目标。 大鼠将暴露于 0.2-0.8 ppm O3 或 5-25 ppm NO2。 治疗方法 细胞骨架不稳定（秋水仙碱和细胞松弛素 B）或稳定 （鬼笔环肽和激动素）药物将单独或组合使用 与污染物暴露相结合。 粘膜通透性的变化 将使用同位素追踪大分子（摩尔重量 469 至 69,000） 该实验室引入了大约两个标签程序 几年前，现在已经很成熟了。 分子运输动力学 将在不同的实验条件下进行比较。 通过光学和电子显微镜进行的免疫细胞化学将用于 表征各种细胞骨架成分及其与 通过紧密连接或内吞囊泡运输。 冷冻骨折 复制品将进行形态测量分析，以检测紧密的细微变化 药物治疗后的连接深度和膜内链或 污染物暴露。 上皮细胞表面形态、损伤 细胞膜和细胞间隙的形成 购买扫描电子将监测相邻细胞的收缩 显微镜。

The proposed study will investigate a) mechanisms which control airway permeability, b) intracellular sites of early injury following inhalation exposure of rats to 03 and N02 and c) means to block or reverse pollutant induced increased permeability. Pilot studies from our laboratory demonstrate an increase in tracheal and bronchoalveolar permeability to macromolecules following inhalation exposure of rats to 03. The increased permeability appears to result, at least in part, from the perturbation of tight junctions. Based on our preliminary observations on the distribution of actin and myosin in the apical cytoplasm and in close vicinity of tight junctions of ciliated epithelial cells and also based on the similarities in the actions of pollutant gases and cytoskeleton-disrupting drugs, we propose that the cytoskeleton is a likely intracellular target site for pollutant and that it plays a critical role in the modulation of tight junction permeability. We also anticipate the role of cytoskeleton in vesicular transport in airways as in other systems. The proposed studies take into account an extensive comparison of cytoskeleton-active drugs with pollutant gases and possible reversal of pollutant effects by the drugs. A variety of indicators will be analyzed to achieve the projected goals. Rats will be exposed to 0.2-0.8 ppm O3 or 5-25 ppm NO2. Treatments with cytoskeleton-destabilizing (colchicine and cytochalasin B) or stabilizing (Phalloidin and kinetin) drugs will be carried out either alone or in combination with pollutant exposures. Changes in mucosal permeabilities to macromolecules (mol wt 469 to 69,000) will be followed using isotope labeling procedures which were introduced in this laboratory about two years ago and are now well established. Kinetics of molecular transport under various experimental conditions will be compared. Immunocytochemistry by light and electron microscopy will be employed to characterize various cytoskeletal components and their relation to transport through tight junctions or endocytic vesicles. Freeze fracture replicas will be analyzed morphometrically to detect fine changes in tight junctional depth and intramembranous strands following drug treatments or pollutant exposures. Surface morphology of epithelial cells, injuries to cell membranes and formation of intercellular spaces resulting from retraction of adjacent cells will be monitored buy scanning electron microscopy.