MACROMOLECULAR TRANSPORT ACROSS LUNG ALVEOLAR EPITHELIUM

拟议研究的具体目标是调查 大分子跨膜转运的特点及调控 肺泡上皮屏障。我们最近对单向~(14)C-的研究 通过肺泡上皮的白蛋白流量显示出明显的不对称性 放射性示踪剂流动，其中流出肺泡腔的流量约为4 比流入牙槽腔的流量大几倍。它的大小 标记的白蛋白通量也大于小得多的白蛋白通量 多糖，菊粉。这些观察结果使我们假设 有一种特殊的大分子(如白蛋白)运输 肺泡上皮细胞的作用机制。我们将调查是否有可能 大分子跨膜转运的机制和途径 利用两种分离的肺泡上皮屏障模型 原代培养大鼠肺组织II型肺泡细胞单层 培养和完好无损的两栖动物肺。大鼠II型肺泡细胞单层 生长在多孔表面和完整的牛蛙肺上，制成平板 薄片将安装在Ussing室中，并将研究其 大分子传输特性。单向放射性示踪剂 蛋白质和右旋糖苷的通量测量和表观分析 渗透性属性，将被执行。特定运输 有待研究的机制包括肺泡上皮蛋白分解。 示踪剂和大分子的囊泡运输。要测试 大分子通过囊泡运输的假说 途径，我们将研究吸附和内化的动力学 肺泡上皮细胞的大分子，以及 囊泡转运抑制剂的作用。形态调查 的大分子转运途径将被执行利用 超微结构标记物(例如，胶体金吸附的大分子) 用电子显微镜技术进行内吞作用。的影响 电荷密度将通过利用中性、阳离子和 阴离子大分子。接合的可能的刺激效应 含有聚阳离子的蛋白质也将被测定。非特定效果 大分子在小溶质和水通量上的传输将是 研究以进一步确定大宗内吞途径在 肺泡上皮。预计这些研究将提供 关于大分子的特性和调控的信息 通过肺泡上皮屏障运输，并将有助于 阐明哺乳动物肺泡上皮细胞的运输特性和 肺泡上皮屏障在肺泡上皮细胞的形成、形成和发展中的作用 体内肺泡性肺水肿的消退。

The specific objectives of the proposed studies are to investigate the characteristics and regulation of macromolecular transport across the alveolar epithelial barrier. Our recent studies on unidirectional 14 C- albumin fluxes across the alveolar epithelium show marked asymmetry of radiotracer flow, in which flux out of the alveolar space is about four times greater than flux into the alveolar space. The magnitudes of labeled albumin fluxes are also greater than that for the much smaller polysaccharide, inulin. These observations led us to postulate that there is a specialized macromolecular (e.g., albumin) transport mechanism in alveolar epithelium. We will investigate possible mechanisms and pathways for the translocation of macromolecules across the alveolar epithelial barrier utilizing two isolated models of the tissue, namely, rat lung type II pneumocyte monolayers in primary culture and intact amphibian lungs. Rat type II pneumocyte monolayers grown on porous surfaces and intact bullfrog lungs prepared as flat sheets will be mounted in Ussing chambers and will be studied for their macromolecular transport characteristics. Unidirectional radiotracer flux measurements of proteins and dextrans, and analysis of apparent permeability properties, will be performed. Specific transport mechanisms to be studied include alveolar epithelial proteolysis of tracers and vesicular transport of macromolecules. To test the hypothesis that macromolecules are translocated via vesicular transport pathways, we will study the kinetics of adsorption and internalization of macromolecules by the alveolar epithelial cells, as well as the effects of vesicular transport inhibitors. Morphological investigation of the macromolecular transport pathways will be performed utilizing ultrastructural markers (e.g., colloidal gold-absorbed macromolecules) for endocytosis with electron microscopic techniques. The influence of charge density will be investigated by utilizing neutral, cationic, and anionic macromolecules. Possible stimulatory effects of conjugation of proteins with polycations will also be determined. Nonspecific effects of macromolecular transport on small solute and water fluxes will be studied to further determine the role of bulk endocytotic pathways in the alveolar epithelium. It is expected that these studies will provide information on the characteristics and regulation of macromolecular transport across the alveolar epithelial barrier, and will help elucidate mammalian alveolar epithelial transport properties and the role of the alveolar epithelial barrier in the prevention, formation, and resolution of alveolar pulmonary edema in vivo.