Mechanisms of proteasomal regulation of fibrosis

描述(由申请人提供)： 摘要博莱霉素是研究肺纤维化的常用模型，在气管内给药后，许多药物或遗传干预措施已被证明可以预防纤维化的发展。这些研究为肺纤维化的发展提供了重要的机制见解，并确认转化生长因子-β(TGF-2)和过氧体激活物激活的受体-γ(PPAR3)都是肺纤维化的重要介质。在我们的初步数据中，我们观察到，在给药8天后给小鼠注射蛋白酶体抑制剂，可以显著减轻肺纤维化。在博莱霉素诱导的硬皮病皮肤纤维化模型中也观察到了类似的保护作用。更多的初步数据表明，蛋白酶体抑制导致PPAR3的丰度和活性增加，PPAR3作为转化生长因子-2的抑制因子发挥作用。我们假设，蛋白酶体抑制剂的应用阻止了泛素介导的PPAR-3在正常人肺成纤维细胞和小鼠肺中的降解，从而抑制了对活性转化生长因子-2的转录反应，从而减轻了纤维化。我们已经产生了三个相互关联的特定目标来确定PPAR3被降解的分子机制以及在转化生长因子-2存在的情况下加速这种降解的分子机制。目的1.在正常肺成纤维细胞中，是否需要PPAR-3来抑制蛋白酶体抑制引起的对活性转化生长因子-2的转录反应？目的2.PPAR-3如何靶向于正常肺成纤维细胞中蛋白酶体的降解？目的3.博来佐米介导的PPAR3蛋白丰度增加是否阻止了博莱霉素治疗的小鼠在转化生长因子-21活化下游发生肺纤维化？这项应用代表着一项高度创新的努力，它利用细胞培养系统中的分子工具和复杂的小鼠模型来阐明蛋白酶体抑制可能阻止肺纤维化发展的机制。我们的初步数据支持拟议实验的可行性，并为我们关注泛素-蛋白酶体系统对PPAR3的调控提供了支持。 公共卫生相关性： 许多肺部疾病，包括急性呼吸窘迫综合征、特发性肺纤维化、结节病、硬皮病和其他各种疾病，会导致肺部的疤痕(纤维化)，如果进展，可能会导致呼吸衰竭或死亡。在这项提案中，研究人员发现，FDA批准的用于治疗某些癌症的药物Bortezomib在预防小鼠肺部和皮肤纤维化方面有效。他们提出了一系列实验，以确定这种药物如何预防纤维化。这些实验的早期数据表明，Bortezomib增加了参与纤维化发展的一种关键蛋白的浓度。靶向这种蛋白将代表着一种治疗退伍军人肺纤维化的新的、新的治疗策略。

DESCRIPTION (provided by applicant): ABSTRACT A number of pharmacologic or genetic interventions have been shown to prevent the development of fibrosis following the intratracheal administration of bleomycin, a commonly used model for the study of lung fibrosis. These studies have provided important mechanistic insights into the development of pulmonary fibrosis and have identified both transforming growth factor-beta (TGF-2) and peroxsome prolifeator- activated receptor-gamma (PPAR3) as important mediators of fibrosis. In our preliminary data, we observed that the administration of a proteasome inhibitor to mice 8 days after the administration of bleomycin resulted in marked attenuation of lung fibrosis. Similar protection was observed in a bleomycin induced skin fibrosis model of scleroderma. Additional preliminary data suggest that proteasomal inhibition results in increased abundance and activity of PPAR3, which functions as an inhibitor of TGF-2. We hypothesize that the administration of proteasomal inhibitors prevents the ubiquitin-mediated degradation of PPAR-3 in normal human lung fibroblasts and in the mouse lung thereby inhibiting the transcriptional response to active TGF-2 and attenuating fibrosis. We have generated three interrelated specific aims to identify the molecular mechanisms by which PPAR3 is degraded and by which this degradation is accelerated in the presence of TGF-2. Aim 1. Is PPAR-3 required for inhibition of the transcriptional response to active TGF-2 induced by proteasomal inhibition in normal human lung fibroblasts? Aim 2. How is PPAR-3 targeted for proteasomal degradation in normal human lung fibroblasts? Aim 3. Does the bortezomib- mediated increase in the protein abundance of PPAR3 prevent the development of lung fibrosis in mice treated with bleomycin downstream of the activation of TGF- 21? This application represents a highly innovative effort that employs molecular tools in cell culture systems and sophisticated mouse models to elucidate the mechanisms by which proteasomal inhibition might prevent the development of pulmonary fibrosis. Our preliminary data support the feasibility of the proposed experiments and provide support for our focus on the ubiquitin-proteasomal system's regulation of PPAR3. PUBLIC HEALTH RELEVANCE: Many lung diseases including the Acute Respiratory Distress Syndrome, Idiopathic Pulmonary Fibrosis, Sarcoidosis, Scleroderma and a variety of others cause scarring (fibrosis) of the lung that if progressive can result in respiratory failure or death. In this proposal, the investigators have found that the administration of an FDA approved medication used to treat some cancers, bortezomib, is effective at preventing the development of lung and skin fibrosis in mice. They propose a series of experiments to determine how this medication protects against fibrosis. Early data from these experiments suggest that bortezomib increases the concentration of a key protein involved in the development of fibrosis. Targeting this protein would represent a new and novel therapeutic strategy for the treatment of Veterans with lung fibrosis.