PPAR gamma Signaling and Lung Fibroblast Transdifferentiation

描述(申请人提供)：异常损伤/修复，伴随肺泡化截断，是“新的支气管肺发育不良(BPD)”的主要组织病理学特征。我们最近提出，破坏正常的内稳态上皮-间充质通讯，以及随之而来的肺泡间质成纤维细胞(AIF)到肌成纤维细胞(MYF)的转分化，是其病理生物学中的关键事件。参与这一过程的具体分子机制仍不完全清楚。本研究的目的是利用体内和体外模型，研究BPD的分子机制，特别是PPAR信号在高氧诱导的大鼠肺AIF-MYF转分化中的作用，并确定一种新的分子预防和治疗方法的有效性。在具体目标1中，我们将利用激光捕获显微解剖、形态计量学、免疫组织化学、实时荧光聚合酶链式反应、Northern和Western分析，在活体新生大鼠模型中确定PPAR激动剂如何预防和/或治疗高氧诱导的AIF到MYF的转分化。针对甲状旁腺激素相关蛋白受体、PPAR、C/增强子结合蛋白、脂肪细胞分化相关蛋白的mRNA表达，以及相应蛋白的差异表达和磷酸化，利用Real Time-PCR、Northern和Western分析、代谢组学、反义和体外转染等方法，探讨AIF向MYF转分化的机制。我们还将确定如何通过罗格列酮和GW7845等有效的PPAR配体刺激造脂途径来防止或逆转AIF到MYF的转分化。这项建议除了为BPD的病理生物学提供了新的见解外，还具有巨大的潜力，可以开辟新的介入策略来治疗一般的慢性肺部疾病，特别是BPD。事实上，使用我们建议中采用的功能基因组方法，即诱导造脂转录因子，不仅可以预防，而且可以逆转已建立的慢性肺部疾病。这项建议中提出的概念是新颖、创新的，与传统的氧致肺损伤范式不同，可能具有比简单理解氧致肺损伤更广泛的含义。

DESCRIPTION (provided by applicant): Abnormal injury/repair, with truncation of alveolarization, is the major histopathological hallmark of "The New Bronchopulmonary Dysplasia (BPD)". We have recently proposed that disrupting normal homeostatic epithelial-mesenchymal communications, and the consequent alveolar interstitial fibroblast (AIF)-to-myofibroblast (MYF) transdifferentiation, are the key events in its pathobiology. The specific molecular mechanisms involved in this process remain incompletely defined. The objective of this proposal is to determine the specific molecular mechanisms involved in BPD, particularly the role of Peroxisome Proliferator Activated Receptor (PPAR) signaling in hyperoxia-induced rat lung AIF-to-MYF transdifferentiation, using both in vivo and in vitro models and to determine the effectiveness of a novel molecular preventive and therapeutic approach. In Specific Aim 1, using Laser Capture Microdissection, Morphometry, Immunohistochemistry, Real Time-PCR, Northern, and Western analyses, we will determine, in an in vivo neonatal rat model, how PPAR agonists prevent and/or treat hyperoxia-induced AIF-to-MYF transdifferentiation. In Specific Aim 2, using Real Time -PCR, Northern and Western analyses, Metabolomics, Antisense, and Transfection in vitro studies, for the mRNA expression of Parathyroid Hormone-related Protein Receptor, PPAR, C/Enhancer Binding Protein, and Adipocyte Differentiation Related Protein, coupled with the differential expression and phosphorylation of the corresponding proteins, we will determine the mechanism involved in AIF-to-MYF transdifferentiation. We will also determine how AIF-to-MYF transdifferentiation can be prevented or reversed by stimulating the lipogenic pathway through potent PPAR ligands such as rosiglitazone and GW7845. This proposal, in addition to providing new insights into the pathobiology of BPD, has enormous potential for opening up novel interventional strategies to tackle chronic lung disease in general, and BPD in particular. In fact, using the functional genomic approach, adopted in our proposal, i.e., inducing lipogenic transcription factors, may not only prevent, but may also reverse established chronic lung disease. The concept put forward in this proposal is novel, innovative, and departs from the traditional paradigm of oxygen-induced lung damage, and may have much wider implications than simply understanding oxygen-induced lung injury.