NEUROPEPTIDES IN LUNG DEVELOPMENT AND INJURY

我们的长期目标是阐明类铃蟾肽的作用， 灵长类动物肺神经内分泌细胞（PNECs）的多肽（BLP） 肺发育，并确定其在治疗 早产儿预防呼吸窘迫综合征（RDS）， 支气管肺发育不良（BPD）。 BLP可能会参与 正常的哺乳动物肺发育，因为高BLP水平被发现， 胎儿肺 我们以前证明了哺乳动物的瞬时表达， BLP mRNA在妊娠中期人胎肺近端到远端的表达 与呼吸道的生长平行。 子宫内BLP 给药加速胎肺生长和成熟;内源性 用抗BLP单克隆抗体阻断肺成熟。 细胞 已发现表面酶CE 10/中性内肽酶24.11（CD 10/NEP 水解和水解BLP，BLP是正常支气管的有丝分裂原， 上皮细胞、肺成纤维细胞和许多小细胞癌 肺的。 SCH 32615增强人胎鼠CD 10/NEP的抑制作用 肺在器官培养物中的生长以及鼠的生长和成熟 子宫内的胎儿肺 上述作用均被BLP受体阻断 对手。 用外源性BLP或 通过抑制CD 10/NEP增强内源性BLP可能预防BPD。 狒狒是唯一可用的动物模型来研究这一重要的 人类失调 我们的第一个目标是描述PNEC个体发育和 BLP、BLP受体和CD 10/NEP基因在发育正常人中的表达 狒狒胎肺和患有肺透明膜病（HMD）的动物， 波士顿警局 其次，我们将确定BLP与 地塞米松（DEX）和/或甲状腺激素（T4）用于灵长类动物胎肺 在器官培养物中生长和成熟，作为体内研究的前奏。 将使用以下方法评估生长：3 H-胸苷掺入细胞核 DNA、PCNA免疫染色和组织病理学。 成熟度将是 用3 H-胆碱掺入到表面活性剂磷脂中进行评估; 组织病理学和电子显微镜检查;免疫染色标记物 分化的II型细胞和PNEC;以及定量相对mRNA 表面活性剂磷脂合成的水平。 第三，我们将调查 BLP与其他药物的最佳组合的体内效应 对狒狒胎肺在妊娠140天的生长和成熟的影响， 将按照上述方式进行评估。 第四，我们将确定 最好的治疗方案显示， 目标3下的生长抑制改善了胎儿的临床结局 狒狒在妊娠140天分娩，我们主要关注BPD。 最后，我们将评估PNEC和BLP，BLP 受体和CE 10/NEP基因在体内发育肺中的表达 在正常狒狒中用Dex加T4或用维甲酸（RA）治疗 以及患有HMD和BPD的动物。 这项调查将是一个关键的 这些药物在临床上的应用的前奏。

Our long-term objectives are to clarify the role of bombesin-like peptides (BLPs) from pulmonary neuroendocrine cells (PNECs) in primate lung development and to determine their clinical usefulness in treating premature infants to prevent respiratory distress syndrome (RDS) and bronchopulmonary dysplasia (BPD). BLPs are likely to participate in normal mammalian lung development because high BLP levels are found in fetal lung. We previously demonstrated transient expression of mammalian BLP mRNAs in mid-gestation human fetal lung in a proximal-to distal fashion in parallel with growth of the airways. In utero BLP administration accelerated fetal lung growth and maturation; endogenous lung maturation was blocked by an anti-BLP monoclonal antibody. the cell surface enzyme CE10/neutral endopeptidase 24.11 (CD10/NEP) has been found to hydrolyze and inactivate BLPs, which are mitogens for normal bronchial epithelial cells, pulmonary fibroblasts, and many small cell carcinomas of the lung. Inhibition of CD10/NEP by SCH32615 potentiated human fetal lung growth in organ cultures and both growth and maturation in murine fetal lung in utero. All of these effects were blocked by BLP receptor antagonists. It is possible that treatment with exogenous BLPs or potentiation of endogenous BLPs by CD10/NEP inhibition might prevent BPD. The baboon is the only available animal model to study this important human disorder. Our First Aim is to characterize PNEC ontogeny and the expression of BLP, BlP receptor, and CD10/NEP genes in developing normal fetal baboon lung and in animals with hyaline membrane disease (HMD) and BPD. Second, we will determine he optimal combination of BLPs with dexamethasone (DEX) and/or thyroid hormone (T4) for primate fetal lung growth and maturation in organ cultures as a prelude to in vivo studies. Growth will be assessed using: 3H-thymidine incorporation into nuclear DNA; PCNA immunostaining; and histopathology. Maturation will be assessed using: 3H-choline incorporation into surfactant phospholipids; histopathology and electron microscopy; immunostaining for markers of differentiated type II cells and PNECs; and quantitation of relative mRNA levels for surfactant phospholipid synthesis. Third, we will investigate in vivo effects of the optimal combination(s) of BLPs with other agents on baboon fetal lung growth and maturation at 140 days gestation, which will be assessed as described above. Fourth, we will determine whether the best treatment regimen shown to enhance lung maturation without growth inhibition under Aim 3 improves the clinical outcome of fetal baboons delivered at 140 days gestation, with our major focus on BPD. Finally, we will assess alterations in PNECs and expression of BLP, BLP receptor, and CE10/NEP genes in developing lung following in vivo treatment with Dex plus T4, or with retinoic acid (RA) in normal baboons and in animals with HMD and BPD. This investigation would be a critical prelude to the application of these agents in clinical medicine.