Role of HIF-1alpha in Fetal Lung Epithelial Differentiation

描述（申请人提供）：肺表面活性剂，由肺泡II型细胞合成分泌，可降低气液界面表面张力。肺表面活性物质对正常肺功能至关重要，这一事实强调了肺表面活性物质缺乏与新生儿呼吸窘迫综合征（RDS）无可争议的联系。在妊娠末期，肺表面活性物质的合成和储存急剧增加。然而，II型细胞成熟的分子调控尚不完全清楚。我们的初步数据表明，缺氧诱导因子-la （HIF-1）在肺上皮分化中起重要作用。肺上皮细胞中cre介导的HIF-1缺失可导致新生儿呼吸衰竭死亡。受感染动物的肺部显示出表面活性剂系统中几个关键成分的显著减少。目前的提议将验证一个假设，即子宫内的缺氧环境对妊娠末期增加表面活性剂的产生至关重要，而HIF-1调节了这一过程的关键方面。我们将在三个特定的目标中研究HIF-1影响肺上皮细胞分化的机制。在第一个特定目标中，我们将使用转基因模型来确定肺特异性HIF-1（缺失）如何调节整体表面活性剂磷脂的生物合成。我们的初步数据还表明，表面活性剂蛋白B （SP-B）和脂质转运蛋白ABCA3的表达在HIF-1缺失的小鼠上皮中降低。在第二个特定目标中，我们将使用肺特异性HIF-1缺失的小鼠来确定HIF-1在调节SP-B和ABCA3中的作用。我们将在体外测试HIF-1直接激活SP-B和ABCA3转录的能力，并确定其与缺氧反应元件的相互作用。我们将使用转基因小鼠模型，其中组成活性形式的HIF-1在肺上皮中诱导表达，以确定HIF-1是否对SP-B和ABCA3表达有直接影响。在第三个特定目标中，我们将在肺上皮中表达一种诱导的显性阴性HIF-1，以确定HIF-1在肺发育的特定时期是否至关重要。我们还将使用该模型来确定有条件表达的SP-B或ABCA3是否可以逆转hif - 1缺失表型。通过确定HIF-1的作用，这些研究将为远端肺上皮分化的调控提供重要的新信息。

DESCRIPTION (provided by applicant): Pulmonary surfactant, which is synthesized and secreted by alveolar type II cells, reduces surface tension at the air-liquid interface. The critical importance of pulmonary surfactant to normal lung function is underscored by the fact that a deficiency in pulmonary surfactant has been incontrovertibly linked to respiratory distress syndrome (RDS) in neonates. Synthesis and storage of pulmonary surfactant increase dramatically at the end of gestation. The molecular regulation of type II cell maturation, however, is not yet completely understood. Our preliminary data demonstrate that hypoxia inducible factor-la (HIF-1() plays an important role in lung epithelial differentiation. Cre-mediated deletion of HIF-1( in lung epithelial cells results in neonatal death from respiratory failure. Lungs from affected animals show significant reductions in several key components of the surfactant system. The present proposal will test the hypothesis that the hypoxic environment in utero is critical for increased surfactant production at the end of gestation, and that HIF-1( regulates key aspects of this process. We will examine the mechanism(s) by which HIF-1( influences lung epithelial cell differentiation in three specific aims. In the first specific aim we will use a transgenic model to determine how lung-specific HIF-1( deletion regulates overall surfactant phospholipid biosynthesis. Our preliminary data also indicate that expression of surfactant protein B (SP-B) and the lipid transporter ABCA3 are decreased in the epithelium of mice in which HIF-1( has been deleted. In the second specific aim we will use mice with lung-specific HIF-1( deletion to determine the role of HIF-1( in the regulation of SP-B and ABCA3. We will test the ability of HIF-1( to directly activate SP-B and ABCA3 transcription in vitro and define its interactions with hypoxia response elements. We will use a transgenic mouse model in which a constitutively active form of HIF-1( is inducibly expressed in the lung epithelium to determine if the effects of HIF-1( on SP-B and ABCA3 expression are direct. In third specific aim, we will express an inducible, dominant-negative form of HIF-1( in the lung epithelium to determine if HIF-1( is critical during particular periods of lung development. We will also use this model to determine if conditionally expressed SP-B or ABCA3 can reverse the HIF-l(-deleted phenotype. By defining the role of HIF-1(, these studies will provide important new information about the regulation of distal lung epithelial differentiation.