REGULATION OF RESPIRATORY TRACT FLUID LAYERS

显微镜检查显示，呼吸道内的液体由两部分组成 层次感。睫毛周围有一层深度不等的粘液毯。 SOL，沐浴纤毛的一层透明的物质。环境污染的来源及治理 人们对这些层次的理解还不完全。然而，在囊性纤维化中，它 据认为，表面上皮细胞的异常离子运输改变了 水合作用这些层之间的关系或水合作用 粘液纤毛清除。在这个提案中，我们将检验两个假设。 首先，我们认为表面上皮细胞的离子转运是 在调节睫毛周围溶胶的深度方面几乎不重要，这是 通过毛细的力量精确地设定在纤毛的长度 由大的组合圆周长度(每平方厘米600米)产生 纤毛的表面)。其次，我们提出了有缺陷的客户关系 而腺体分泌的液体主要负责最初的 囊性纤维化中粘液堆积。这些缺陷导致了 脱水的粘液不会从腺体开口完全分离 和/或不会有效地被纤毛运输。此外 腺体中CFTR的故障可能会改变硫酸盐化或其他特性 腺体粘液，再次损害粘液纤毛清除。为了测试这些 假设，我们将研究渗透和活性液体如何流过 表面上皮改变粘液层的深度，以及这些 变化会影响粘液的运输。流体层将在中可视化 扫描电子显微镜下快速冷冻组织的横切面 显微镜。我们将测量介体诱导的体液分泌跨CF和 使用电容探针技术的非CF细胞培养，并从完整 用微吸管从导管开口取样腺体。我们将确定是否 从CF腺和表面培养物中提取的粘蛋白的硫酸盐化和流变学 不正常。最后，我们将确定本地化和总级别 CFTR在我们的文化中存在，并与激素诱导的 这些参数随功能的变化而变化。其中大部分建议 实验将在我们最近开发的原代培养物上进行 表面和腺体上皮，显示出高水平的超微结构 和功能分化。

Microscopy has shown the fluid lining the airways to consist of two layers. A mucous blanket of variable depth lies on top of the periciliary sol, a clear layer which bathes the cilia. The sources and regulation of these layers are incompletely understood. However, in cystic fibrosis, it is believed that abnormal ion transport by the surface epithelium alters the relationship or hydration of these layers in such a way as to impair mucociliary clearance. In this proposal we will test two hypotheses. Firstly, we suggest that ion transport by the surface epithelium is of little importance in regulating the depth of the periciliary sol, which is set precisely at the length of the cilia by forces of capillarity generated by the large combined circumferential length (600 m per cm2 of epithelial surface) of the cilia. Secondly, we propose that defective Cl and fluid secretion by the glands is mainly responsible for the initial accumulation of mucus in cystic fibrosis. These defects result in dehydrated mucus which will not detach fully from the gland openings and/or will not be effectively transported by the cilia. In addition malfunction of CFTR in glands may alter the sulfation or other properties of gland mucus, again impairing mucociliary clearance. To test these hypotheses, we will investigate how osmotic and active fluid flows across the surface epithelium alter the depth of the mucus layers, and how these changes affect mucus transport. The fluid layers will be visualized in transverse sections of rapidly frozen tissues in the Scanning electron microscope. We will measure mediator-induced fluid secretion across CF and non-CF cell cultures using a capacitance probe technique, and from intact glands by micropipette sampling from duct openings. We will determine if the sulfation and rheology of mucins from CF gland and surface cultures is abnormal. Finally, we will determine the localization and total levels of CFTR in our cultures, and correlate hormonally-induced alterations in these parameters with changes in function. Most of these proposed experiments will be performed on our recently developed primary cultures of surface and gland epithelia, which show high levels of ultrastructural and functional differentiation.