TRANSFORMING GROWTH FACTOR - BETA MEDIATES EFFECTS OF HYPOXIA IN NEWBORN LUNG

描述（由申请方提供）：慢性缺氧导致发育中肺的异常肺动脉重塑（PAR）和肺泡发育抑制（IAD）。转化生长因子-β（TGF-β）是一种肽类生长因子，是肺发育和血管重塑的关键调节因子，可能是PAR和IAD发病机制中的重要介质。TGF-β作为无活性前体合成，并且TGF-β的活化是引发生物学效应所必需的关键步骤。TGF-β活化被肺中表达细胞表面糖蛋白Thy-1的成纤维细胞抑制。我们最近已经表明，由于诱导型显性负性TGF-β II型受体（DN 2 RII）而导致TGF-β信号传导中断的新生小鼠减弱了IAD和PAR，表明TGF-β信号传导在IAD和PAR中是重要的。我们已经证明，缺氧减少Thy-1和增加活性TGF-β在新生小鼠，表明缺氧诱导的Thy-1的减少可能允许增加TGF-β的激活。我们的实验室已经进行了新的观察，Thy-1缺失小鼠具有IAD和增加的磷酸化Smad 2、β-平滑肌肌动蛋白、间质胶原、组织肺阻力（但正常气道阻力）和降低的肺顺应性，证明在发育中的肺中缺乏Thy-1导致类似于支气管肺发育不良（BPD）的肺表型。我们的初步数据表明，TGF-β的合成可能是由内皮素-1（ET-1），一个已知的调解PAR。本项目的目的是确定肺发育过程中低氧暴露导致TGF-β激活增加以及随后的PAR和IAD的机制。具体目标1将检验TGF-β信号传导对于缺氧诱导的PAR和IAD是必需的这一假设。具体目标2将评估Thy-1在TGF-β 1活化中的机制作用以及在缺氧存在或不存在的情况下IAD的发病机制。具体目标3将检验缺氧诱导的ET-1增加刺激TGF-β合成和活化的具体假设。将使用垂直整合方法，包括体内模型（新生小鼠从出生起暴露于缺氧或空气2周）和体外模型（新生鼠肺动脉平滑肌细胞和成纤维细胞）。除野生型小鼠外，目前可用的转基因小鼠（Thy-1 null，DN 2 RII，（6整合素null））将用于表征关键信号传导途径。PAR可导致新生儿持续性肺动脉高压（PPHN），而IAD和PAR均见于BPD。尽管新生儿护理取得了进展，但PPHN和BPD的发病率仍然很高，新的范例对于开发新的治疗策略至关重要。在这个建议中概述的实验将增加我们的知识的机制负责正常和异常的肺动脉重塑和肺泡发育在围产期。至少，这些研究将确定TGF-β作为低氧诱导的IAD和PAR的关键调节因子的作用。这些实验还将鉴定由缺氧调节的TGF-β信号通路的组分和缺氧条件下的TGF-β激活剂。项目叙述：新生儿死亡和发病的两个主要原因是新生儿持续性肺动脉高压（PPHN）和支气管肺发育不良（BPD），其特征在于异常肺动脉增厚、肺动脉高压和肺泡发育抑制。转化生长因子-β（一种在正常肺发育中很重要的蛋白质）的过度活化可能导致PPHN和BPD中观察到的异常肺发育。该提案将确定转化生长因子-β在异常肺发育中的作用以及转化生长因子-β被激活的机制，并可能导致PPHN和BPD新治疗方案的开发。

DESCRIPTION (provided by applicant): Chronic hypoxia leads to abnormal pulmonary arterial remodeling (PAR) and inhibition of alveolar development (IAD) in the developing lung. Transforming growth factor-beta (TGF-(), a peptide growth factor that is a key regulator of lung development and vascular remodeling, may be a crucial mediator in the pathogenesis of PAR and IAD. TGF-( is synthesized as an inactive precursor, and activation of TGF-( is the critical step necessary to elicit biological effects. TGF-( activation is inhibited by fibroblasts in the lung which express the cell surface glycoprotein Thy-1. We have recently shown that newborn mice with disruption of TGF-( signaling due to inducible dominant negative TGF-( type II receptors (DN2RII) have attenuated IAD and PAR, indicating that TGF-( signaling is important in IAD and PAR. We have shown that hypoxia reduces Thy-1 and increases active TGF-( in newborn mice, suggesting that hypoxia-induced decreases in Thy-1 may permit increased TGF-( activation. Our laboratory has made the novel observations that Thy-1 null mice have IAD and increased phospho-Smad2, (-smooth muscle actin, interstitial collagen, tissue lung resistance (but normal airway resistance), and decreased lung compliance, demonstrating that absence of Thy-1 in the developing lung leads to a lung phenotype similar to bronchopulmonary dysplasia (BPD). Our preliminary data indicate that TGF-( synthesis may be regulated by endothelin-1 (ET-1), a known mediator of PAR. The objective of this project is to determine the mechanisms by which hypoxic exposure during lung development leads to increased TGF-( activation and subsequent PAR and IAD. Specific Aim 1 will test the hypothesis that TGF-( signaling is necessary for hypoxia-induced PAR and IAD. Specific Aim 2 will assess the mechanistic role of Thy-1 in TGF-( activation and the pathogenesis of IAD in the presence or absence of hypoxia. Specific Aim 3 will test the specific hypothesis that hypoxia-induced increases in ET-1 stimulate TGF-( synthesis and activation. A vertically-integrated approach, with in vivo models (newborn mice exposed to hypoxia or air for 2 weeks from birth) and in vitro models (neonatal murine pulmonary arterial smooth muscle cells and fibroblasts) will be used. Currently available transgenic mice (Thy-1 null, DN2RII, (6 integrin null), in addition to wild-type mice will be used to characterize the critical signaling pathways. PAR contributes to persistent pulmonary hypertension of the newborn (PPHN), and both IAD and PAR are seen in BPD. The incidence of PPHN and BPD remains high despite advances in neonatal care, and new paradigms are essential for the development of novel therapeutic strategies. The experiments outlined in this proposal will increase our knowledge of the mechanisms responsible for normal and abnormal pulmonary arterial remodeling and alveolar development in the perinatal period. At a minimum, these studies will determine the role of TGF-( as a key regulator of hypoxia-induced IAD and PAR. These experiments will also identify the components of TGF-( signaling pathway regulated by hypoxia, and TGF-( activators under hypoxic conditions. PROJECT NARRATIVE: Two major causes of death and morbidity in newborn infants are persistent pulmonary hypertension of the newborn (PPHN) and bronchopulmonary dysplasia (BPD), which are characterized by abnormal pulmonary arterial thickening, pulmonary hypertension, and inhibition of alveolar development. Excessive activation of transforming growth factor-beta, a protein that is important in normal lung development, may lead to the abnormal lung development that is seen in PPHN and BPD. This proposal will determine the role of transforming growth factor-beta in abnormal lung development and the mechanisms by which transforming growth factor-beta becomes activated, and may lead to the development of new treatment options for PPHN and BPD.