The Role of Mitochondrial Reactive Oxygen Species in TGF-Beta Signaling

许多药理学或遗传干预已被证明可以预防纤维化的发展 在肺内施用博来霉素（一种用于肺纤维化研究的常用模型）之后。 这些研究为肺纤维化的发展提供了重要的机制见解， 已经确定了转化生长因子-β（TGF-β）和过氧化物酶体增殖剂激活的受体- γ（PPAR-g）作为纤维化的重要介质。我们已经证明，防止过氧化物酶体增殖物激活受体的降解- g对TGF-β的反应损害正常人肺中胶原和其他促纤维化基因的表达 成纤维细胞，来自肺纤维化患者的肺成纤维细胞，来自肺纤维化患者的皮肤成纤维细胞， 硬皮病和用博来霉素治疗的小鼠。此外，我们已经生成了初步数据， TGF-β诱导线粒体活性氧（ROS），这有助于降解 PPAR-g和是完整的TGF-β转录反应所必需的。我们假设TGF-β诱导的 线粒体ROS通过Smad 3激活通过ALK 5受体。这些衍生的ROS 激活下游激酶，诱导PPAR-g降解，放大TGF-β的表达 依赖基因我们建议在一系列体外实验中，在动物模型中， 急性肺损伤/纤维化和来自肺纤维化和ARDS患者的肺泡液。目标1： 确定TGF-β诱导线粒体ROS产生的机制。目标2：确定 线粒体ROS调节TGF-β诱导的基因转录的机制。目标3：确定 成纤维细胞特异性线粒体ROS是否是小鼠肺纤维化发展所必需的 模型和线粒体活性氧在肺结核患者TGF-β介导的基因表达中的重要性 纤维化该应用代表了在细胞培养中采用分子工具的高度创新的努力 系统和复杂的小鼠模型，以阐明线粒体ROS调节的机制， 肺纤维化的发展。我们的初步数据支持建议的可行性 实验，并提供支持，我们的重点是TGF-β介导的调节PPAR-g通过诱导 线粒体ROS。我们的研究小组的组成和所有拟议的实验设计 以鉴定用于治疗肺纤维化的新的治疗靶点。

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-b) and peroxsome prolifeator-activated receptor- gamma (PPAR-g) as important mediators of fibrosis. we have shown that preventing the degradation of PPAR- g in response to TGF-¿ impairs the expression of collagen and other profibrotic genes in normal human lung fibroblasts, lung fibroblasts from patients with pulmonary fibrosis, skin fibroblasts from patients with scleroderma and mice treated with bleomycin. In addition, we have generated preliminary data suggesting that TGF-b induces mitochondrial reactive oxygen species (ROS), which contribute to the degradation of PPAR-g and are required for a full TGF-b transcriptional response. We hypothesize that TGF-b-induced mitochondrial ROS via Smad3 activation through the ALK5 receptor. These mitochondrially derived ROS activate downstream kinases and induce the degradation of PPAR-g to amplify the expression of TGF-b dependent genes. We propose to test these hypotheses in a series of experiments in vitro, in animal models of acute lung injury/fibrosis and with alveolar fluid from patients with pulmonary fibrosis and ARDS. Aim-1: To determine the mechanism by which TGF-¿ induces the generation of mitochondrial ROS. Aim-2: To determine the mechanism by which mitochondrial ROS modulate TGF-¿-induced gene transcription. Aim-3: To determine whether fibroblast specific mitochondrial ROS are required for the development of lung fibrosis in murine models and the importance of mitochondrial ROS in TGF-¿-mediated gene expression in patients with lung fibrosis. 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 mitochondrial ROS regulate the development of pulmonary fibrosis. Our preliminary data support the feasibility of the proposed experiments and provide support for our focus on the TGF-b mediated regulation of PPAR-g via induction of mitochondrial ROS. The composition of our research group and all of the proposed experiments are designed to identify novel therapeutic targets for the treatment of lung fibrosis.