LUNG ELASTIN IN BRONCHOPULMONARY DYSPLASIA

支气管肺发育不良中的肺弹性蛋白

• 批准号: 6390507
• 负责人: Richard A Pierce
• 金额: $ 19.35万
• 依托单位: BARNES-JEWISH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-01 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6390507
• 关键词: RNase protection assay; amine oxidoreductase; baboons; bronchopulmonary dysplasia; cooperative study; disease /disorder model; elastases; elastin; enzyme activity; enzyme induction /repression; fibrillin; genetic transcription; immunocytochemistry; in situ hybridization; lung; messenger RNA; premature infant animal; protein biosynthesis

The focus of this proposal is to characterize mechanisms controlling elastin synthesis and turnover during the development of bronchopulmonary dysplasia (BPD) resulting from ventilation of the premature lung. Elastin confers the requisite property of elastic recoil to such lung structures as alveoli and alveolar ducts, bronchioles, and blood vessels, and thus is essential for lung function. We have previously demonstrated abnormal elastic fiber deposition in an experimental model of BPD, and others have demonstrated increased elastolytic activity in the BPD lung. Still, the root causes of elastic fiber abnormalities in BPD are not known. We hypothesize that mesenchymal cells of the premature lung respond to the strain of mechanical ventilation by increasing the expression and deposition of elastic extracellular matrix components out of proportion to what is required for alveolarization. Exposure to hyperoxia may result in the production and release of elastases such as neutrophil elastase or matrix metalloproteinases that damage elastic fibers, as well as cytokines or growth factors that alter extracellular matrix gene expression by lung fibroblasts. These events result in the excess deposition of disordered elastic fibers at sites of failed development of new alveolar walls. The accumulation of disorganized elastic fibers at these sites may limit the ability to recover from injury. To determine the causes of abnormal elastic fiber deposition during the development of BPD, we propose to study the expression of elastin, fibrillins 1 and 2, and lysyl oxidase, all required for normal elastic fiber synthesis, and to characterize the elastases present in the injured lung. Our experimental approaches will include quantitative analysis of elastic fiber-related and elastase mRNA expression by RNAse protection assays, as well as localization of expression by in situ hybridization and immunohistochemical analyses. The molecular mechanisms regulating tropoelastin expression during normal baboon lung development and the development of BPD will be determined by assessing changes in tropoelastin gene transcription, steady-state mRNA levels, and protein synthesis, expression and activity of elastases in the BPD lung will be assessed by a combination of substrate zymography, immunohistochemistry, and in situ hybridization. Determining the effects of interventive treatments on the expression of elastic fiber-related genes and the elaboration of elastases will test the hypothesis that treatments which prevent BPD will also restore normal patterns of elastic fiber-related gene expression and elastic fiber deposition.

本提案的重点是表征在支气管肺发育不良（BPD）的发展过程中控制弹性蛋白合成和转换的机制。弹性蛋白赋予肺泡和肺泡管、细支气管和血管等肺结构弹性反冲的必要性质，因此对肺功能至关重要。我们之前在BPD的实验模型中证实了异常的弹性纤维沉积，其他人也证实了BPD肺的弹性分解活性增加。然而，BPD中弹性纤维异常的根本原因尚不清楚。我们假设，早产儿肺的间充质细胞对机械通气压力的反应是增加弹性细胞外基质成分的表达和沉积，与肺泡化所需的比例不成比例。暴露于高氧环境可能导致弹性蛋白酶的产生和释放，如中性粒细胞弹性蛋白酶或基质金属蛋白酶，它们会损伤弹性纤维，以及细胞因子或生长因子，它们会改变肺成纤维细胞的细胞外基质基因表达。这些事件导致无序弹性纤维在新肺泡壁发育失败的部位过量沉积。无序弹性纤维在这些部位的积累可能会限制损伤恢复的能力。为了确定BPD发展过程中异常弹性纤维沉积的原因，我们建议研究弹性蛋白、纤维蛋白1和2以及赖氨酸氧化酶的表达，这些都是正常弹性纤维合成所必需的，并表征损伤肺中存在的弹性酶。我们的实验方法将包括通过RNAse保护实验定量分析弹性纤维相关和弹性酶mRNA表达，以及通过原位杂交和免疫组织化学分析定位表达。在正常狒狒肺发育和BPD发展过程中，调节弹力蛋白表达的分子机制将通过评估弹力蛋白基因转录、稳态mRNA水平和蛋白质合成的变化来确定，BPD肺中弹性蛋白酶的表达和活性将通过底物酶谱、免疫组织化学和原位杂交的组合来评估。确定介入治疗对弹性纤维相关基因表达和弹性蛋白酶的影响，将检验预防BPD的治疗也将恢复弹性纤维相关基因表达和弹性纤维沉积的正常模式的假设。