LUNG EPITHELIAL ION-LIQUID TRANSPORT DURING DEVELOPMENT

这项提议的目的是阐明离子传输特性如何 在发育过程中呼吸道上皮细胞的变化 哺乳动物出生前的一个充满液体的、分泌氯离子的器官的肺 出生后充满空气、吸收钠的器官。总体战略是 为了确定细胞内离子转运的发育变化 呼吸道上皮的2个不同区域(气管、远端 2种(兔、羊)，并将这些变化与 出生前后肺液体平衡的发育差异。 长期目标是提高对基本机制的理解 导致流经肺上皮细胞的液体倒流 出生，这应该为发展有效的 抑制或促进婴儿肺水肿消退的措施， 无论是孩子还是成年人。具体目标侧重于以下中心假设 与分娩和出生适应有关的事件，特别是 释放肾上腺激素(皮质醇、醛固酮和肾上腺素)， 影响上皮细胞代谢和Na-K-ATPase活性， 结果是从分泌氯离子到吸收钠离子的转换 支气管肺上皮。该项目使用4个互补的 钠和氯的作用机制及其调控的实验研究方法 发育中动物呼吸道上皮的转运：(1)呼吸道 远端肺上皮细胞将从胎儿、新生儿和 两种成年动物的放射性标记通量的测量 离子(~(86)Rb、~(36)Cl、~(22)Na)在存在或不存在各种抑制剂的情况下 离子输运特性；(2)生物电学和离子输运特性 培养的呼吸道和远端肺上皮细胞融合单层 将在细胞事先暴露于特定环境和不接触特定环境的情况下进行评估 激素(糖皮质激素、醛固酮、β-肾上腺素能激动剂) 妊娠后期浓度增加；(3)净生产量 将测量腔内液体的(分泌和吸收)。 胎儿和新生动物(兔， 在基础和实验条件下(激素暴露， 使用刺激剂和上皮离子转运抑制剂进行治疗)； 和(4)净液体产量、离子浓度和跨上皮 将测量胎羊肺的电位差 在分娩前和分娩期间，事先接触和不接触激素，以及 在存在或不存在影响上皮离子的各种药物的情况下 运输。这些互补的实验方法允许(A) 不同地区和地区离子输运性质的评估 发育中的呼吸道上皮的不同细胞；和(B) 澄清它们各自对液体分泌的贡献和 活体肺在相关生理条件下的吸收 动物。这项工作可能对流体有重要的影响 新生儿肺的平衡和与呼吸障碍相关的 呼吸道上皮功能障碍。

The purpose of this proposal is to clarify how ion transport properties of the respiratory epithelium change during development to convert the mammalian lung from a liquid-filled, Cl-secreting organ before birth to an air-filled, Na-absorbing organ after birth. The general strategy is to define developmental changes of ion transport in cells derived from 2 distinct regions of the respiratory tract epithelium (trachea, distal lung) of 2 species (rabbits, sheep), and to relate these changes to developmental differences in lung liquid balance before and after birth. The longterm objective is to improve understanding of basic mechanisms responsible for reversal of liquid flow across the lung epithelium near birth, which should provide useful information for developing effective measures to inhibit or promote resolution of pulmonary edema in infants, children and adults. Specific aims focus on the central hypothesis that events associated with labor and adaptation at birth, in particular the release of adrenal hormones (cortisol, aldosterone, and epinephrine), influence epithelial cell metabolism and Na-K-ATPase activity, with a resultant switch from Cl-secretion to Na-absorption across the bronchopulmonary epithelium. The project uses 4 complementary experimental approaches to study mechanisms and regulation of Na and Cl transport in the respiratory epithelium of developing animals: (1) airway and distal lung epithelial cells will be isolated from fetal, newborn and adult animals of 2 species in order to measure fluxes of radiolabeled ions (86Rb,36Cl, 22Na) in the presence or absence of various inhibitors of ion transport; (2) bioelectric and ion transport properties of confluent monolayers of cultured airway and distal lung epithelial cells will be assessed with and without prior exposure of the cells to specific hormones (glucocorticoids, aldosterone, beta-adrenergic agonists) that increase in concentration late in gestation; (3) net production (secretion and absorption) of luminal liquid will be measured in isolated, blood-perfused lungs of fetal and newborn animals (rabbits, sheep) under basal and experimental conditions (hormone exposure, treatment with stimulants and inhibitors of epithelial ion transport); and (4) net liquid production, ion concentrations and transepithelial electrical potential difference will be measured in lungs of fetal sheep before and during labor, with and without prior exposure to hormones, and in the presence or absence of various drugs that influence epithelial ion transport. These complementary experimental approaches permit (a) assessment of ion transport properties at different regions and in different cells of the developing respiratory epithelium; and (b) clarification of their respective contributions to liquid secretion and absorption under relevant physiological conditions in lungs of living animals. This work may have important implications concerning fluid balance in the newborn lung and breathing disorders associated with respiratory epithelial dysfunction.