INSULIN AND HUMAN FETAL LUNG DEVELOPMENT

描述：此修订后的申请建议研究影响， 胰岛素对人胎肺表面活性蛋白A的作用机制 糖尿病母亲的胎儿患新生儿肺的风险增加 疾病，并且已经提出高水平的胰岛素延迟人类的 肺成熟申请人先前的研究表明， 胰岛素处理人胎肺外植体不影响表面活性剂 磷脂合成或组成，但确实减少了 表面活性物质蛋白A及其mRNA。此外， 糖尿病患者的羊水低于非糖尿病患者 妇女在这项建议中，假设高胰岛素血症可能 糖尿病母亲的婴儿易患呼吸窘迫 通过抑制表面活性剂相关蛋白的合成， 特别是SP-A，从而改变了 表面活性提出了使用培养的人胎肺进行实验， H-441腺癌细胞系，探讨胰岛素作用机制 方面的影响.第一个目标将确定胰岛素调节的两个 人SP-A基因在培养组织中，并使用原位杂交， 将两种SP-A转录本定位在人肺组织中。第二 Specific Aim将研究胰岛素调节的分子机制 SP-A mRNA稳定性研究，以及 抑制剂的第三个具体目标将调查3'的作用。 SP-A mRNA的非翻译部分，其被预测为介导 胰岛素的抑制作用。瞬时转染研究， 嵌合基因将确定参与拟议的 胰岛素的不稳定作用。在第四个具体目标中， 参与胰岛素与糖皮质激素相互作用的基因 将检测表达，包括转录速率、转录物 稳定性和可能诱导新的基因产物， PCR法这些研究将提供关于其作用的新信息， 胰岛素的作用机制及其与糖皮质激素的相互作用 人胎肺发育

DESCRIPTION: This revised application proposes to study effects and mechanisms of insulin action on surfactant protein A of human fetal lung. The fetus of the diabetic mother is at increased risk for newborn lung disease and it has been proposed that high levels of insulin delay human lung maturation. The previous studies by the applicant indicated that insulin treatment of human fetal lung explants did not affect surfactant phospholipid synthesis or composition but did decrease accumulation of surfactant protein A and its mRNA. Also, concentrations of SP-A in amniotic fluid were found to be lower in diabetic vs. non-diabetic women. In this proposal it is hypothesized that hyperinsulinemia may predispose infants of diabetic mothers to develop respiratory distress by inhibiting the synthesis of surfactant associated proteins, in particular SP-A, thereby altering the composition and function of surfactant. Experiments are proposed using cultured human fetal lung and the H-441 adenocarcinoma cell line to explore the mechanism of insulin effects. The first aim will determine insulin regulation of the two human SP-A genes in cultured tissue and use in situ hybridization to localize the two SP-A transcripts in human lung tissue. The second Specific Aim will examine molecular mechanisms for insulin regulation of SP-A using nuclear run-on assays, studies of SP-A mRNA stability, and inhibitors. The third specific aim will investigate the role of 3' untranslated portion of SP-A mRNA which is predicted to mediate the inhibitory effects of insulin. Transient transfection studies with chimeric genes will identify sequences involved in the proposed destabilizing effect of insulin. In the fourth specific aim, mechanisms involved in the interaction between insulin and glucocorticoid in gene expression will be examined including transcription rate, transcript stability and possible induction of new gene products using differential PCR. These studies will provide new information on the role and mechanism of insulin effects and interaction with glucocorticoids in human fetal lung development.